Stabilized Modified Release Vitamin D Formulation and Method of Administering Same

ABSTRACT

A stabilized formulation for controlled release of a vitamin D compound is disclosed. The formulation comprises one or both of 25-hydroxyvitamin D2 and 25-hydroxyvitamin D3 and a cellulosic compound. The stabilized formulations exhibit a stable dissolution profile following exposure to storage conditions and demonstrate improved pharmacokinetic parameters compared to unstabilized formulations.

CROSS-REFERENCE TO RELATED APPLICATION

This is a continuation of U.S. patent application Ser. No. 15/720,948,filed Sep. 29, 2017, which is a continuation of application Ser. No.14/213,285, filed Mar. 14, 2014, now U.S. Pat. No. 9,861,644 issued Jan.9, 2018, which claims the benefit of priority under 35 U.S.C. § 119(e)of U.S. Provisional Patent Application No. 61/801,896, filed Mar. 15,2013, the priority benefits of which are hereby claimed and thedisclosures of which are incorporated herein by reference.

BACKGROUND Field of the Disclosure

The disclosure relates generally to controlled release pharmaceuticalcompositions. More particularly, the invention relates tocontrolled-release formulations for delivery of a vitamin D compound forintestinal absorption, such as a 25-hydroxyvitamin D compound, which areshelf-stable over time.

Brief Description of Related Technology

The Vitamin D metabolites known as 25-hydroxyvitamin D₂ and25-hydroxyvitamin D₃ (collectively referred to as “25-hydroxyvitamin D”)are fat-soluble steroid prohormones that contribute to the maintenanceof normal levels of calcium and phosphorus in the bloodstream. Theprohormone 25-hydroxyvitamin D₂ is produced from Vitamin D₂(ergocalciferol) and 25-hydroxyvitamin D₃ is produced from Vitamin D₃(cholecalciferol), primarily by one or more enzymes located in theliver. The two prohormones also can be produced outside of the liverfrom Vitamin D₂ and Vitamin D₃ (collectively referred to as “Vitamin D”)in certain cells, such as enterocytes, which contain enzymes identicalor similar to those found in the liver.

The 25-hydroxyvitamin D prohormones are further metabolized in thekidneys into potent Vitamin D hormones. The prohormone 25-hydroxyvitaminD₂ is metabolized into 1α,25-dihydroxyvitamin D₂; likewise,25-hydroxyvitamin D₃ is metabolized into 1α,25-dihydroxyvitamin D₃(calcitriol). Production of these active hormones from the25-hydroxyvitamin D prohormones also can occur outside of the kidney incells which contain the required enzyme(s).

Controlled release formulations of 25-hydroxyvitamin D₂ and/or25-hydroxyvitamin D₃ can be administered to treat 25-hydroxyvitamin Dinsufficiency and deficiency without supraphysiological surges inintraluminal, intracellular and blood levels of 25-hydroxyvitamin D andtheir consequences; without causing substantially increased catabolismof the administered 25-hydroxyvitamin D; and, without causing seriousside effects associated with Vitamin D supplementation, namely Vitamin Dtoxicity. The controlled release formulations effectively lower PTHlevels without undesirable increases in serum calcium and serumphosphorus and are therefore useful for treating secondaryhyperparathyroidism, for example in CKD patients. See InternationalPatent Application Nos. PCT/US2007/061521 and PCT/US2008/061579 and U.S.patent application Ser. No. 12/109,983, incorporated herein byreference.

The controlled released compositions provide substantially increasedabsorption of 25-hydroxyvitamin D via transport on DBP and decreasedabsorption via transport in chylomicrons. The compositions also providemaintenance of substantially constant blood levels of 25-hydroxyvitaminD during a 24-hour post-dosing period. By providing a gradual, sustainedand direct release of 25-hydroxyvitamin D₂/25-hydroxyvitamin D₃ andabsorption preferentially to circulating DBP (rather than tochylomicrons), blood, intraluminal and intracellular 25-hydroxyvitamin Dconcentration spikes, i.e., supraphysiologic levels and related unwantedcatabolism can be mitigated or eliminated. Furthermore, by providing agradual and sustained release, serum levels of 25-hydroxyvitamin D canbe increased and maintained more predictably than by administration ofimmediate release formulations, allowing for a consistent dosage andreducing or eliminating the need for frequent patient monitoring.

To deliver the benefits of controlled release formulations of25-hydroxyvitamin D to patients, there is a need for stabilizedpharmaceutical compositions that retain the desired dissolutionproperties of the formulation, for extended periods of time, e.g., aftershipping and storage.

SUMMARY

The present invention comprises a controlled release Vitamin Dformulation comprising a vitamin D compound and a cellulosic compound.

The present invention also comprises a storage-stabilized formulationfor controlled release of a vitamin D compound in the gastrointestinaltract of a subject which ingests the formulation. In one aspect, thestabilized formulation comprises one or both of 25-hydroxyvitamin D₂ and25-hydroxyvitamin D₃ and a stabilizing agent or stabilizing compound,e.g. a cellulosic compound. The stabilized formulations of the inventionwith the recited stabilizing agent(s) can have improved or relativelyimproved “storage stability”, or stability following aging, as well asone or more additional characteristics including improved physical,chemical and biological properties when compared to the disclosedformulations that do not contain such agents. The claimed formulationsare thus suitable as therapeutics that possess a long shelf life as wellas improved bioavailability compared to aged, unstable formulations.

In one embodiment, the stabilized formulation includes one or both of25-hydroxyvitamin D₂ and 25-hydroxyvitamin D₃, a lipophilic matrix (e.g.a wax matrix), and a stabilizing agent (e.g., a cellulosic compound). Inone aspect, a stabilized formulation includes one or both of25-hydroxyvitamin D₂ and 25-hydroxyvitamin D₃, a wax matrix, and acellulosic stabilizing agent. In another aspect, the formulationincludes one or both of 25-hydroxyvitamin D₂ and 25-hydroxyvitamin D₃, awax matrix, and an effective amount of a cellulosic compound to maintainan advantageous degree of stabilization described herein.

In one type of embodiment, the stabilized formulation comprises amixture of an active-loaded wax matrix comprising one or both of25-hydroxyvitamin D₂ and 25-hydroxyvitamin D₃ and a cellulosicstabilizing agent, wherein the formulation releases an amount of25-hydroxyvitamin D during in vitro dissolution after exposure tostorage conditions of at least one month at 25° C. and 60% relativehumidity that varies at all dissolution testing time points by 30% orless compared to the amount released at the same dissolution time pointsduring in vitro dissolution conducted on fresh product.

In one type of embodiment, the formulation is an improvement formulationfor controlled release of a vitamin D compound. In one aspect, theimprovement comprises admixing a stabilizing agent into a formulationfor controlled release of a vitamin D compound in the gastrointestinaltract of a subject which ingests the formulation. In another aspect, theimprovement comprises an effective amount of a cellulosic compoundadmixed into a formulation for controlled release of a vitamin Dcompound in the gastrointestinal tract of a subject which ingests theformulation to provide an advantageous degree of stability describedherein.

In one embodiment, the invention comprises a stable sustained releasevitamin D formulation comprising 25-hydroxyvitamin D₂ or25-hydroxyvitamin D₃ or combinations thereof and sustained releaseexcipients wherein said formulation has a dissolution profile X at T₀that retains this profile according to the formula X=T₀+/−30% overstorage conditions selected from room temperature and ambient humidity,or 25° C. and 60% RH, or 40° C. and 75% RH, for example.

Further aspects and advantages will be apparent to those of ordinaryskill in the art from a review of the following detailed description,taken in conjunction with the drawings. While the compositions andmethods are susceptible of embodiments in various forms, the descriptionhereafter includes specific embodiments with the understanding that thedisclosure is illustrative, and is not intended to limit the inventionto the specific embodiments described herein.

For the compositions and methods described herein, optional features,including but not limited to components, compositional ranges thereof,substituents, conditions, and steps, are contemplated to be selectedfrom the various aspects, embodiments, and examples provided herein.

BRIEF DESCRIPTION OF THE DRAWINGS

FIGS. 1A, 1B, and 1C show the dissolution profile of formulationsaccording to the disclosure after storage for 0 to 24 months at 25° C.and 60% relative humidity. The dissolution time in hours is depicted onthe x-axis and the mean percent of 25-hydroxyvitamin D₃ dissolved isshown on the y-axis. FIGS. 1A, 1B, and 1C show the dissolution profileof formulations comprising 30 μg, 60 μg, and 90 μg of 25-hydroxyvitaminD₃, respectively.

FIGS. 2A, 2B, and 2C show the dissolution profile of formulationsaccording to the disclosure after storage for 0 to 6 months at 40° C.and 75% relative humidity. The dissolution time in hours is depicted onthe x-axis and the mean percent of 25-hydroxyvitamin D₃ dissolved isshown on the y-axis. FIGS. 2A, 2B, and 2C show the dissolution profileof formulations comprising 30 μg, 60 μg, and 90 μg of 25-hydroxyvitaminD₃, respectively.

FIGS. 3A and 3B show the dissolution profile of formulations afterstorage for 0 to 12 months at 25° C. and 60% relative humidity. Thedissolution time in hours is depicted on the x-axis and the mean % LabelClaim of 25-hydroxyvitamin D₃ released is shown on the y-axis. FIG. 3Ashows the dissolution profile of a comparative formulation that does notcontain a cellulosic compound. FIG. 3B shows the dissolution profile ofa stabilized formulation according to the disclosure.

FIG. 4 shows the resulting mean baseline-adjusted calcifediolconcentrations by treatment group (PK population) for the patientsdescribed in Example 4 treated with a formulation according to thedisclosure.

FIG. 5 shows the resulting summary baseline-adjusted PK parameters forcalcifediol concentrations by treatment group (PK population) for thepatients described in Example 4 treated with a formulation according tothe disclosure.

FIG. 6 shows the resulting mean baseline-adjusted serum1,25-dihydroxyvitamin D levels during the 6-week treatment (PKpopulation) for the patients described in Example 4 treated with aformulation according to the disclosure.

FIG. 7 shows a summary of the resulting baseline-adjusted repeat-dose PKparameters for serum 1,25-dihydroxyvitamin D by treatment group (PKpopulation) for the patients described in Example 4 treated with aformulation according to the disclosure.

FIG. 8 shows the resulting mean percent of baseline in plasma iPTHlevels during the 6-week treatment (PK population) for the patientsdescribed in Example 4 treated with a formulation according to thedisclosure.

FIG. 9 shows a summary of the resulting baseline-adjusted repeat-dose PKparameters for plasma iPTH by treatment group (PK population) for thepatients described in Example 4 treated with a formulation according tothe disclosure.

FIGS. 10 and 11 show the percent change from baseline at EOT for plasmaiPTH relative to baseline-adjusted calcifediol and 1,25-dihydroxyvitaminD exposure (AUC_(0-6wk)) in the PK Population for the patients describedin Example 4 treated with a formulation according to the disclosure.

DETAILED DESCRIPTION

As used herein, the terms “controlled release,” and “modified release”are used interchangeably and refer to the release of the administeredvitamin D compound in a way that deviates from immediate release. Asused herein, the terms “sustained release” and “extended release” areused interchangeably and refer to the release of the administeredvitamin D compound over a longer period of time than a comparableimmediate release formulation, resulting in serum concentrations of thevitamin D compound that remain elevated over baseline for a longerperiod of time than for a comparable immediate release formulation. Theforegoing terms optionally include delayed release characteristics. Forexample, a delayed release type of controlled release formulation willbe characterized by Cmax at a time greater than Cmax for an immediaterelease formulation. As another example, the release of a25-hydroxyvitamin D compound will preferably be at such a rate thattotal serum or blood levels of 25-hydroxyvitamin D are maintained orelevated above predosing levels for an extended period of time, e.g. 4to 24 hours or even longer.

As used herein, the term “cellulosic compound” can include cellulose(C₆H₁₀O₅)_(n) or a derivative of cellulose, unless specified otherwise.A “cellulose ether” is a cellulose derivative that has been chemicallymodified to result in partial or complete etherification of the hydroxylgroups in the cellulose molecule. Examples of cellulose derivativeswhich can be used as stabilizing agents include, but are not limited to,celluloronic acid, carboxy methyl cellulose, ethyl cellulose,hydroxyethyl cellulose, hydroxyl propyl cellulose, hydroxyl propylmethylcellulose, methyl cellulose, polyanionic cellulose, andcombinations thereof, for example. Different grades of each cellulosiccompound or stabilizing agent, corresponding to variations in, e.g.,molecular weight, viscosity, solubility, and hydration, are alsoencompassed by the terms.

Any vitamin D compound suitable for prophylactic and/or therapeutic use,and combinations thereof, are contemplated for inclusion in theformulation described herein. Vitamin D, 25-hydroxyvitamin D,1,25-dihydroxyvitamin D, and other metabolites and analogs of Vitamin Dare also useful as active compounds in pharmaceutical compositions.Specific examples include, but are not limited to, Vitamin D₃(cholecalciferol), Vitamin D₂ (ergocalciferol), 25-hydroxyvitamin D₃,25-hydroxyvitamin D₂, 25-hydroxyvitamin D₄, 25-hydroxyvitamin D₅,25-hydroxyvitamin D₇, 1α,25-dihydroxyvitamin D₃, 1α,25-dihydroxyvitaminD₂, 1α,25-dihydroxyvitamin D₄, and vitamin D analogs (including allhydroxy and dihydroxy forms), including 1,25-dihydroxy-19-nor-vitaminD₂, and 1α-hydroxyvitamin D₃. In one type of embodiment, the vitamin Dcompound includes one or more hydroxy forms, such as a combination of25-hydroxyvitamin D₃ and 25-hydroxyvitamin D₂.

A type of vitamin D compound particularly contemplated for use in theformulation disclosed herein can include 25-hydroxyvitamin D₂,25-hydroxyvitamin D₃, or a combination thereof. 25-hydroxyvitamin D₃, isparticularly contemplated. As used herein, the term 25-hydroxyvitamin Drefers to one or more of 25-hydroxyvitamin D₃, 25-hydroxyvitamin D₂,25-hydroxyvitamin D₄, 25-hydroxyvitamin D₅, or 25-hydroxyvitamin D₇, andit is contemplated that in any reference thereto a preferred embodimentis one or more of 25-hydroxyvitamin D₃ and 25-hydroxyvitamin D₂,preferably 25-hydroxyvitamin D₃. Thus, in any and all formulationsdescribed herein, it is specifically contemplated that the active caninclude one or both of 25-hydroxyvitamin D₂ and 25-hydroxyvitamin D₃,particularly 25-hydroxyvitamin D₃. In the disclosure herein, the vitaminD compound (or combination thereof) is also referred to as the “active”part of the formulation (or “active” agents), as distinguished from thecontrolled release matrix, the stabilizing agent, and other excipients.In the pharmacokinetic testing reported herein with samples that used25-hydroxyvitamin D₃ as the active, references to 25-hydroxyvitamin Dshould be interpreted to mean 25-hydroxyvitamin D₃, and allpharmacokinetic (PK) results associated with (e.g., t_(max), C_(max),AUC) should be understood to be based on 25-hydroxyvitamin D₃.

As used herein, a “stabilized” formulation refers to a formulationexhibiting a stable in vitro dissolution profile (according to any ofthe parameters described further herein) and controlled release (e.g.,sustained release) of a vitamin D compound in vivo, for a time followinginitial manufacture, e.g. following actual shelf storage or acceleratedstability storage conditions. The release of the active ingredient canbe measured using a suitable in vitro dissolution method, such as one ofthe methods already known in the art. In principle, any of thedissolution studies described in the United States Pharmacopeia, USP29-NF 24, Dissolution <711> physical tests and determinations, UnitedStates Pharmacopeial Convention, Inc., Rockville, Md., 2006, pp.2673-2682.; European Pharmacopoeia 2.9.3 Dissolution Test for SolidDosage Forms, or the Japanese Pharmacopoeia 6.10 Dissolution Test, canbe used to determine if a formulation is stable. For purposes of thepresent invention, the in vitro dissolution method is United StatesPharmacopeia, USP 29-NF 24, Dissolution <711> physical tests anddeterminations, United States Pharmacopeial Convention, Inc., Rockville,Md., 2006, pp. 2673-2682, using Apparatus 2 (paddle method), asdescribed in the Examples below.

As used herein, t_(max) (or Tmax) is defined as the time for the plasmaconcentration of the active compound to reach its maximum in a doseinterval following administration of a formulation according to theinvention. When administering a single 25-hydroxyvitamin D compound, forexample 25-hydroxyvitamin D₃, tmax is defined as the time for the plasmaconcentration of serum 25-hydroxyvitamin D₃ to reach its maximum in adose interval following administration of the formulation, unlessspecified otherwise.

Consistent with the NKF K/DOQI Guidelines, as used herein Vitamin Dsufficiency is defined as serum 25-hydroxyvitamin D levels ≥30 ng/mL,Vitamin D insufficiency is defined as serum 25-hydroxyvitamin D of 16-30ng/mL, mild Vitamin D deficiency is defined as serum 25-hydroxyvitamin Dof 5-15 ng/mL, and severe Vitamin D deficiency is defined as serum25-hydroxyvitamin D below 5 ng/mL.

In jurisdictions that forbid the patenting of methods that are practicedon the human body, the meaning of “administering” of a composition to ahuman subject shall be restricted to prescribing a controlled substancethat a human subject will self-administer by any technique (e.g.,orally, inhalation, topical application, injection, insertion, etc.).The broadest reasonable interpretation that is consistent with laws orregulations defining patentable subject matter is intended. Injurisdictions that do not forbid the patenting of methods that arepracticed on the human body, the “administering” of compositionsincludes both methods practiced on the human body and also the foregoingactivities.

It is specifically understood that any numerical value recited hereinincludes all values from the lower value to the upper value, i.e., allpossible combinations of numerical values between the lowest value andthe highest value enumerated are to be considered to be expressly statedin this application. For example, if a concentration range or abeneficial effect range is stated as 1% to 50%, it is intended thatvalues such as 2% to 40%, 10% to 30%, or 1% to 3%, etc., are expresslyenumerated in this specification. As another example, a statedconcentration of about 20% is intended to include values from 19.5% upto 20.5%. These are only examples of what is specifically intended.

Disclosed herein are formulations for controlled release of a Vitamin Dcompound in the gastrointestinal tract of a subject which ingests theformulation. The formulation will include a vitamin D compound asdescribed herein, a matrix component that releasably binds the vitamin Dcompound and controllably releases the vitamin D compound (e.g., alipophilic matrix), and a stabilizer (e.g. a cellulosic compound).

A stabilized formulation according to the disclosure herein, followingstorage for a period of time, releases an amount of 25-hydroxyvitamin Din in vitro dissolution that does not substantially differ from thedissolution of the same formulation just after manufacturing and priorto storage. For example, in one embodiment, a formulation releases anamount of 25-hydroxyvitamin D during in vitro dissolution after exposureto storage conditions of two months at 25° C. and 60% relative humiditythat varies at any given dissolution time point after four hours by 30%dr less compared to the amount released at the same dissolution timepoint during in vitro dissolution conducted prior to exposing theformulation to the storage conditions (i.e., freshly-produced product).

The table below provides examples of advantageous degrees of storagestability contemplated for embodiments of the invention followingstorage at 25° C. and 60% RH, and alternatively at 40° C. and 75% RH forvarious times following initial manufacturing, and at various times induring dissolution testing. The degrees of storage stability areexpressed in terms of the maximum deviation from nominal active potency,i.e. maximum % change from LC. Alternative embodiments of maximumdeviation are also provided.

Time (h) 1 month 3 mos. 6 mos. 9 mos. 12 mos. 18 mos. 24 mos. storage at25° C. and 60% RH 2 30%, or 25%, 30%, or 30%, or 30%, or 30%, or 30%, or30%, or or 20%, or 25%, or 25%, or 25%, or 25%, or 25%, or 25%, or 15%,or 10% 20%, or 20%, or 20%, or 20%, or 20%, or 20%, or 15%, or 15%, or15%, or 15%, or 15%, or 15%, or 10% 10% 10% 10% 10% 10% 4 30%, or 25%,30%, or 30%, or 30%, or 30%, or 30%, or 30%, or or 20%, or 25%, or 25%,or 25%, or 25%, or 25%, or 25%, or 15%, or 10% 20%, or 20%, or 20%, or20%, or 20%, or 20%, or 15%, or 15%, or 15%, or 15%, or 15%, or 15%, or10% 10% 10% 10% 10% 10% 6 30%, or 25%, 30%, or 30%, or 30%, or 30%, or30%, or 30%, or or 20%, or 25%, or 25%, or 25%, or 25%, or 25%, or 25%,or 15%, or 10% 20%, or 20%, or 20%, or 20%, or 20%, or 20%, or 15%, or15%, or 15%, or 15%, or 15%, or 15%, or 10% 10% 10% 10% 10% 10% 8 30%,or 25%, 30%, or 30%, or 30%, or 30%, or 30%, or 30%, or or 20%, or 25%,or 25%, or 25%, or 25%, or 25%, or 25%, or 15%, or 10% 20%, or 20%, or20%, or 20%, or 20%, or 20%, or 15%, or 15%, or 15%, or 15%, or 15%, or15%, or 10% 10% 10% 10% 10% 10% 12 30%, or 25%, 30%, or 30%, or 30%, or30%, or 30%, or 30%, or or 20%, or 25%, or 25%, or 25%, or 25%, or 25%,or 25%, or 15%, or 10% 20%, or 20%, or 20%, or 20%, or 20%, or 20%, or15%, or 15%, or 15%, or 15%, or 15%, or 15%, or 10% 10% 10% 10% 10% 10%storage at 40° C. and 75% RH 2 30%, or 25%, 30%, or 30%, or 30%, or 30%,or 30%, or 30%, or or 20%, or 25%, or 25%, or 25%, or 25%, or 25%, or25%, or 15%, or 10% 20%, or 20%, or 20%, or 20%, or 20%, or 20%, or 15%,or 15%, or 15%, or 15%, or 15%, or 15%, or 10% 10% 10% 10% 10% 10% 430%, or 25%, 30%, or 30%, or 30%, or 30%, or 30%, or 30%, or or 20%, or25%, or 25%, or 25%, or 25%, or 25%, or 25%, or 15%, or 10% 20%, or 20%,or 20%, or 20%, or 20%, or 20%, or 15%, or 15%, or 15%, or 15%, or 15%,or 15%, or 10% 10% 10% 10% 10% 10% 6 30%, or 25%, 30%, or 30%, or 30%,or 30%, or 30%, or 30%, or or 20%, or 25%, or 25%, or 25%, or 25%, or25%, or 25%, or 15%, or 10% 20%, or 20%, or 20%, or 20%, or 20%, or 20%,or 15%, or 15%, or 15%, or 15%, or 15%, or 15%, or 10% 10% 10% 10% 10%10% 8 30%, or 25%, 30%, or 30%, or 30%, or 30%, or 30%, or 30%, or or20%, or 25%, or 25%, or 25%, or 25%, or 25%, or 25%, or 15%, or 10% 20%,or 20%, or 20%, or 20%, or 20%, or 20%, or 15%, or 15%, or 15%, or 15%,or 15%, or 15%, or 10% 10% 10% 10% 10% 10% 12 30%, or 25%, 30%, or 30%,or 30%, or 30%, or 30%, or 30%, or or 20%, or 25%, or 25%, or 25%, or25%, or 25%, or 25%, or 15%, or 10% 20%, or 20%, or 20%, or 20%, or 20%,or 20%, or 15%, or 15%, or 15%, or 15%, or 15%, or 15%, or 10% 10% 10%10% 10% 10%

In one type of embodiment, the formulation will have advantageousdegrees of stability described in the table immediately above atmultiple time points throughout the dissolution testing, e.g. at leastat both 2 and 4 hour time points, optionally also at the 6 hour timepoint, further optionally also at the 8 hour time point, and furtheroptionally also at the 12 hour time point, such that the dissolutionprofile after storage follows the dissolution profile of fresh product.Alternatively, the formulation will have advantageous degrees ofstability described in the table immediately above at least at the 2, 6,and 12 hour time points. Alternatively, the formulation will haveadvantageous degrees of stability described in the table immediatelyabove at least at the 4, 8, and 12 hour time points. Alternatively, theformulation will have advantageous degrees of stability described in thetable immediately above at least at the 2, 4, and 6, hour time points.Alternatively, the formulation will have advantageous degrees ofstability described in the table immediately above at least at the 4, 6,8, and 12 hour time points, or at all times of 4 hours and thereafter.

In any and all of the embodiments described in the table immediatelyabove, it is contemplated that the deviation can be positive (morerelease) or negative (less release) with respect to the fresh product.In one type of embodiment, it is contemplated that the deviation will bein the negative (less release) direction at multiple time points. Stillfurther, in one type of embodiment it is contemplated that the deviationin dissolution release would have been negative (less release) atmultiple time points but for the presence of the stabilizing agent inthe formulation.

In any of the embodiments contemplated herein, the dissolution releaseprofile of the formulation can have the characteristics of any one ofthe examples provided herein below. For example, the formulation can becharacterized by a dissolution release profile providing a release ofvitamin D compound of less than 30% at 2 hours, greater than 45% at 6hours, and greater than 80% at 12 hours, and further optionally lessthan 60% at 6 hours.

In another type of embodiment, the formulation can be characterized byan in vitro dissolution profile providing release of vitamin D compoundof less than 30% at 100 to 140 minutes, greater than 45% at 5 to 7hours, and greater than 80% at 11 to 13 hours. In another type ofembodiment, the formulation can be characterized by an in vitrodissolution profile providing release of vitamin D compound of less than30% at 2 hours, greater than 45% at 6 hours, and greater than 80% at 12hours. In these types of embodiments, optionally the release of vitaminD compound at 5 to 7 hours is less than 60%, or at 6 hours is less than60%.

In another type of embodiment, the formulation can be characterized byan in vitro dissolution profile providing release of vitamin D compoundof about 20% to about 40% at 2 hours, at least 35% at 6 hours, and atleast 70% at 12 hours. In another type of embodiment, the formulationcan be characterized by an in vitro dissolution profile providingrelease of vitamin D compound of about 25% to about 35% at 2 hours, atleast 40% at 6 hours, and at least 75% at 12 hours. In these types ofembodiments, optionally the release of vitamin D compound is 75% or lessat 6 hours, or 65% or less at 6 hours, or 60% or less at 6 hours, forexample.

In any of the embodiments described herein, the stabilized formulationcan be characterized by a t_(max) following administration of the dosageform to a human patient, of at least 4 hours, or at least 8 hours, or atleast 12 hours, or at least 18 hours, or at least 20 hours, or at least24 hours, or at least 28 hours, for example in a range of 4 to 96 hours,or in a range of 18 to 30 hours, or in a range of 13 to 28 hours, or is28 hours, for example.

In any of the embodiments contemplated herein a formulation comprising25-hydroxyvitamin D can be characterized by providing abaseline-adjusted C_(max) per microgram of 25-hydroxyvitamin D in arange of about 0.0133 ng/mL to about 0.04 ng/mL when administered to anadult human.

In any of the methods contemplated herein, the method can includeadministering a stabilized sustained release dosage form comprising a25-hydroxyvitamin D compound to a human patient, comprisingadministering an effective amount of the formulation to the patient toprovide a baseline-adjusted C_(max) of at least about 0.2 ng/mL andoptionally less than 110 ng/mL, and further optionally 24 ng/mL or less,for example in a range of about 0.2 to about 24 ng/mL.

In any of the methods contemplated herein, the method can includeadministering a stabilized sustained release dosage form comprising a25-hydroxyvitamin D compound to a human patient, comprisingadministering an effective amount of the formulation to the patient toprovide a baseline-adjusted AUC_(0-inf) of at least 52 ng*h/mL, andoptionally less than 34,500 ng*h/mL, and further optionally about 12,000ng*h/mL or less, for example in a range of about 52 ng*h/mL to about12,000 ng*h/mL.

In any of the embodiments described herein, it is contemplated that thestabilized formulation, following storage, can be bioequivalent to thefreshly-made product. Thus, for example, the stabilized formulation,following storage, can provide an area under the curve for the active(or serum total 25-hydroxyvitamin D), AUC (e.g., AUC_(0-inf) orAUC_(0-t)) within a 90% confidence interval, or within 80% to 125% ofthe mean, or within 80% to 120% of the mean of the fresh product. Inaddition or in the alternative, the stabilized formulation, followingstorage, can provide a maximum serum concentration of the active (orserum total 25-hydroxyvitamin D), C_(max) (e.g., C_(max) absolute, orC_(max) compared to baseline) within a 90% confidence interval, orwithin 80% to 125% of the mean, or within 80% to 120% of the mean, ofthe fresh product.

In one embodiment, a stabilized formulation comprises one or both of25-hydroxyvitamin D₂ and 25-hydroxyvitamin D₃, a wax matrix, and acellulosic compound. In one aspect, a stabilized formulation comprisesone or both of 25-hydroxyvitamin D₂ and 25-hydroxyvitamin D₃, a waxmatrix, and a cellulosic stabilizing agent. In another aspect, theformulation comprises one or both of 25-hydroxyvitamin D₂ and25-hydroxyvitamin D₃, a wax matrix, and an effective amount of acellulosic compound to provide an advantageous degree of stability asdescribed herein, e.g. with respect to the table immediately above orconsistent with any of the Examples described below. For example, theamount can be effective to provide a difference of 30% or less betweenthe amount of active released during in vitro dissolution after exposureto storage conditions of at least one month at 25° C. and 60% relativehumidity at a dissolution time point and the amount released at the samedissolution time point during in vitro dissolution conducted prior toexposing the formulation to the storage conditions, while a comparativeformulation lacking the stabilizing agent would result in a greaterdifference in dissolution release following the same storage conditions.

In one aspect, the formulation is an improved formulation for controlledrelease of a vitamin D compound in the gastrointestinal tract of asubject which ingests the formulation. In one embodiment, theimprovement comprises admixing a cellulosic stabilizing agent into aformulation for controlled release of a vitamin D compound in thegastrointestinal tract of a subject which ingests the formulation. Inanother embodiment, the improvement comprises an effective amount of acellulosic compound admixed into a formulation for controlled release ofa vitamin D compound in the gastrointestinal tract of a subject whichingests the formulation to provide an advantageous degree of stabilityas described herein, e.g. with respect to the table immediately above orconsistent with any of the Examples described below. For example, theamount can be effective to provide a difference of 30% or less betweenthe amount of active released during in vitro dissolution after exposureto storage conditions of at least one month at 25° C. and 60% relativehumidity at a dissolution time point and the amount released at the samedissolution time point during in vitro dissolution conducted prior toexposing the formulation to the storage conditions, while a comparativeformulation lacking the stabilizing agent would result in a greaterdifference in dissolution release following the same storage conditions.

The stabilizing agents can include cellulose compounds. Examples ofcellulose compounds and stabilizing agents for use in the stabilizedformulations of the disclosure can include, but are not limited to,celluloronic acid, carboxy methyl cellulose, ethyl cellulose, hydroxylethyl cellulose, hydroxyl propyl cellulose, hydroxyl propyl methylcellulose, methylcellulose, polyanionic cellulose, and combinationsthereof. Also contemplated are one or more of poloxamers (e.g.,polaxamer 407), poly (ethylene oxide) polymers (e.g., Dow's POLYOXpolymers), povidones, and fumed silicas (e.g., AEROSIL 200, EvonikIndustries AG, Essen, Germany). The stabilizer, e.g. a cellulosiccompound, preferably is present in an amount of at least about 5% of theformulation, based on the total weight of the formulation excluding anyadditional coatings or shells (wt %). For example, the cellulosiccompound can be present in an amount of at least 5 wt % of theformulation, or at least 10 wt % of the formulation, or at least 15 wt %of the formulation, or greater than 5 wt % of the formulation, orgreater than 10 wt % of the formulation, or greater than 15 wt % of theformulation. Suitable ranges include 5 wt % to 30 wt %, 10 wt % to 20 wt%, 10 wt % to 15 wt %, 5 wt % to 15 wt %, and 7.5 wt % to 12.5 wt. %.Examples include about 5 wt %, about 6 wt %, about 7 wt %, about 8 wt %,about 9 wt %, about 10 wt %, about 11 wt %, about 12 wt %, about 13 wt%, about 14 wt %, and about 15 wt %. It will be understood that thestabilizing agent referred to herein is an agent that stabilizes thedissolution release profile (and thus also the in vivo release profile)against substantial change over time during storage conditions, e.g.typical shelf storage conditions. Other agents which are known in theart as preservatives for preventing degradation of the active componentitself are not intended to be encompassed within the terms “stabilizingagent” and “stabilizer” although such preservatives are alsocontemplated for use in the formulations of the present invention.

In one class of embodiment, the cellulosic compound is a celluloseether. Examples of cellulose ethers include, but are not limited to,methylcellulose, hydroxyl propyl methylcellulose, hydroxyl ethylmethylcellulose, hydroxyl ethyl cellulose, hydroxyl propyl cellulose,and combinations thereof.

Hydroxyl propyl methylcellulose (HPMC) is particularly contemplated. TheHPMC can be characterized by one or more of the following features,which are specifically contemplated individually and in combinations.The % methyoxyl component in the HPMC can be in a range of 19 to 24. The% hydroxypropyl component can be in a range of 7 to 12. The apparentviscosity (2% solution in water at 20° C.) can be at least 50,000 cP, orat least 80,000 cP, or in a range of about 80 to 120,000 cP, or 3000 to120,000 cP, or 11,000 to 120,000 cP, or 80,000 to 120,000 cP.Particularly, the apparent viscosity (2% solution in water at 20° C.)can be in a range of 80,000 to 120,000 cP. The pH (1% solution in water)can be in a range of 5.5 to 8.0. For example, a suitable hydroxyl propylmethylcellulose having all of the foregoing properties, including anapparent viscosity (2% solution in water at 20° C.) in a range of 80,000to 120,000 cP, is METHOCEL K100M CR (Dow Wolff Cellulosics, Midland,Mich.).

In one type of embodiment, the cellulosic compound will be insoluble inthe matrix formulation at the melt point of the primary components ofthe matrix, e.g., at 65° C. or in a range of 60° C. to 75° C.

In one type of embodiment, the cellulosic compound will be hydrophilic.

The pharmaceutical formulations according to the disclosure comprisingone of more of 25-hydroxyvitamin D₂ and 25-hydroxyvitamin D₃ and acellulosic compound have unexpectedly improved stability compared toformulations lacking a cellulosic compound. In one embodiment, astabilized formulation according to the disclosure comprises a mixtureof an active-loaded lipophilic matrix comprising one or both of25-hydroxyvitamin D₂ and 25-hydroxyvitamin D₃ and a cellulosicstabilizing agent, wherein the formulation releases an amount of25-hydroxyvitamin D during in vitro dissolution after exposure tostorage conditions of at least one month at 25° C. and 60% relativehumidity that varies at any given dissolution time point by 30% or lesscompared to the amount released at the same dissolution time pointduring in vitro dissolution conducted on freshly-made product.

Formulations that are not stabilized exhibit changes in the amount ofactive ingredient released after the composition is stored for a periodof time, as shown in the Examples below. An unstabilized formulationreleases an amount of 25-hydroxyvitamin D following exposure to storageconditions that can vary at a given dissolution time point, for exampleby more than 30% compared to the amount released at the same dissolutiontime point during in vitro dissolution conducted on freshly-madeproduct. The changes may be an increase or decrease in the dissolutionrate at a given time point, and such changes produce a dissolutionprofile whose curve is distinct from the shape of the initialdissolution profile. An unstabilized formulation also exhibits differentin vivo effects compared to a stabilized formulation according to thedisclosure, following storage as described herein, e.g. following 3months or more of storage at 25° C. and 60% RH. A stabilized formulationdemonstrates different clinical pharmacokinetic parameters, such asimproved bioavailability, compared to an unstabilized formulation,following storage as described herein, e.g. following 3 months or moreof storage at 25° C. and 60% RH. A stabilized formulation according tothe disclosure can have a base formulation which is storage unstable,combined with a stabilizing agent which renders the formulation storagestable as described herein.

The matrix that releasably binds and controllably releases the activecomponent can be, for example, a lipophilic matrix, including a waxmatrix. A wax matrix can provide a formulation which is solid orsemi-solid at room temperature and solid, semi-solid, or liquid at bodytemperature, preferably semi-solid or liquid at body temperature. In oneaspect, the wax matrix comprises a controlled release agent, anemulsifier, and an absorption enhancer.

Examples of controlled release agents suitable for use include, but arenot limited to, waxes, including synthetic waxes, microcrystalline wax,paraffin wax, carnauba wax, and beeswax; polyethoxylated castor oilderivatives, hydrogenated vegetable oils, glyceryl mono-, di- ortribehenates; long-chain alcohols, such as stearyl alcohol, cetylalcohol, and polyethylene glycol; and mixtures of any of the foregoing.Non-digestible waxy substances, such as hard paraffin wax, arepreferred.

The controlled release agent can be present in an amount of at least 5wt % of the formulation, or greater than about 5 wt % of theformulation. For example, depending on the controlled release agentused, the controlled release agent can comprise at least 5 wt % of theformulation or at least 10 wt % of the formulation, or at least 15 wt %of the formulation, or at least 20 wt % of the formulation, or at least25 wt % of the formulation, or greater than 5 wt % of the formulation,or greater than 10 wt % of the formulation, or greater than 15 wt % ofthe formulation, or greater than 20 wt % of the formulation, and orgreater than 25 wt % of the formulation. The controlled release agentcan be present in an amount 50 wt % or less, 40 wt % or less, 35 wt % orless, or 30 wt % or less. Suitable ranges include 5 wt % to 40 wt %, 10wt % to 30 wt % and 15 wt % to 25 wt %. Examples include about 15 wt %,about 16 wt %, about 17 wt %, about 18 wt %, about 19 wt %, about 20 wt%, about 21 wt %, about 22 wt %, about 23 wt %, about 24 wt %, and about25 wt %.

Examples of emulsifiers suitable for use in the formulation include, butare not limited to, lipophilic agents having an HLB of less than 7, suchas mixed fatty acid monoglycerides; mixed fatty acid diglycerides;mixtures of fatty acid mono- and diglycerides; lipophilic polyglycerolesters; glycerol esters including glyceryl monooleate, glyceryldioleate, glyceryl monostearate, glyceryl distearate, glycerylmonopalmitate, and glyceryl dipalmitate; glyceryl-lacto esters of fattyacids; propylene glycol esters including propylene glycol monopalmitate,propylene glycol monostearate, and propylene glycol monooleate; sorbitanesters including sorbitan monostearate, sorbitan sesquioleate; fattyacids and their soaps including stearic acid, palmitic acid, and oleicacid; and mixtures thereof glyceryl monooleate, glyceryl dioleate,glyceryl monostearate, glyceryl distearate, glyceryl monopalmitate, andglyceryl dipalmitate; glyceryl-lacto esters of fatty acids; propyleneglycol esters including propylene glycol monopalmitate, propylene glycolmonostearate, and propylene glycol monooleate; sorbitan esters includingsorbitan monostearate, sorbitan sesquioleate; fatty acids and theirsoaps including stearic acid, palmitic acid, and oleic acid; andmixtures thereof.

A preferred lipoidic agent is selected from glycerides and derivativesthereof. Preferred glycerides are selected from the group consisting ofmedium or long chain glycerides, caprylocaproyl macrogolglycerides, andmixtures thereof.

Preferred medium chain glycerides include, but are not limited to,medium chain monoglycerides, medium chain diglycerides, caprylic/caprictriglyceride, glyceryl monolaurate, glyceryl monostearate,caprylic/capric glycerides, glycerylmonocaprylate, glycerylmonodicaprylate, caprylic/capric linoleic triglyceride, andcaprylic/capric/succinic triglyceride.

Monoglycerides having a low melting point are preferred for making theformulation. Preferred monoglycerides include but are not limited to,glyceryl monostearate, glyceryl monopalmitate, glyceryl monooleate,glyceryl monocaprylate, glyceryl monocaprate, glyceryl monolaurate,etc., preferably glycerol monostearate (GMS). GMS is a naturalemulsifying agent. It is oil soluble, but poorly soluble in water. GMShas an HLB value of 3.8. The lipophilic emulsifier can be present in anamount in a range of about 10 wt % to about 40 wt %, or about 20 wt % toabout 25 wt %, for example. Other examples include about 20 wt %, about21 wt %, about 22 wt %, about 23 wt %, about 24 wt %, and about 25 wt %.

Examples of suitable absorption enhancers include, but are not limitedto, caprylocaproyl macrogolglycerides such as polyethylene glycosylatedglycerides, also known as polyglycolized glycerides or PEGylatedglycerides. PEGylated glycerides which may be employed in thecomposition include, but are not limited to, mixtures of monoglycerides,diglycerides, and triglycerides and monoesters and diesters ofpolyethylene glycol, polyethylene glycosylated almond glycerides,polyethylene glycosylated corn glycerides, and polyethylene glycosylatedcaprylic/capric triglyceride. The absorption enhancer can have an HLBvalue from 13 to 18, or from 13 to 15.

One preferred absorption enhancer is known under the trade name GELUCIRE(Gattefossé Corporation, Paramus, N.J., USA). GELUCIRE is a well knownexcipient which is a family of fatty acid esters of glycerol and PEGesters, also known as polyglycolized glycerides. GELUCIRE is used invarious applications including preparing sustained releasepharmaceutical compositions. GELUCIRE compounds are inert, semi-solidwaxy materials which are amphiphilic and are available with varyingphysical characteristics such as melting point, HLB, and solubilities invarious solvents. They are surface active in nature and disperse orsolubilize in aqueous media forming micelles, microscopic globules orvesicles. They are identified by their melting point/HLB value. Themelting point is expressed in degrees Celsius. One or a mixture ofdifferent grades of GELUCIRE excipient may be chosen to achieve thedesired characteristics of melting point and/or HLB value. A preferredGELUCIRE composition is GELUCIRE 44/14, a mixture of lauroylmacrogolglycerides and lauroyl polyoxylglycerides that has a meltingpoint of 44° C. and a HLB of 14. The absorption enhancer can be presentin an amount of about 5 wt % to about 20 wt %, or about 8 wt % to about15 wt %, for example. Other examples include about 8 wt %, about 9 wt %,about 10 wt %, about 11, wt % about 12 wt %, about 13 wt %, about 14 wt%, and about 15 wt %.

The low melting points of the wax matrix provide a means ofincorporating the pharmaceutically active ingredients, e.g. the vitaminD compound such as 25-D₂, 25-D₃, or both, at temperatures from about 0°C. to about 50° C. above the melting point of the wax matrix and thenfilling the melt (solution and/or dispersion) in suitable capsules. Thecapsules can be of any variety that is compatible with the temperatureof the melt fill, including soft or hard gelatin capsules, and animal orvegetable gelatin capsules. The melt solidifies inside the capsules uponcooling to room temperature.

In one aspect, a stabilized formulation may further comprise an oilyvehicle for the 25-hydroxyvitamin D₂ and/or 25-hydroxyvitamin D₃. Anypharmaceutically-acceptable oil can be used. Examples include animal(e.g., fish), vegetable (e.g., soybean), and mineral oils. The oilpreferably will readily dissolve the 25-hydroxyvitamin D compound used.Preferred oily vehicles include non-digestible oils, such as mineraloils, particularly liquid paraffins, and squalene. The oily vehicle canbe present at a concentration in a range about 10 wt % to about 50 wt %of the formulation, or about 15 wt % to about 45 wt %, or about 20 wt %to about 40 wt %, or about 30 wt % to about 40 wt %, for example. In onetype of embodiment, a suitable liquid paraffin can be characterized byone or more of the following parameters: specific gravity about 0.88 to0.89; kinematic viscosity (40° C.) about 64 cSt to about 70 cSt;molecular weight 424; % paraffinic hydrocarbons about 59; and pour point−24° C. The ratio between the wax matrix and the oily vehicle can beoptimized in order to achieve the desired rate of release of the vitaminD compound. Thus, if a heavier oil component is used, relatively less ofthe wax matrix can be used, and if a lighter oil component is used, thenrelatively more wax matrix can be used.

The stabilized controlled release compositions in accordance with theinvention preferably are designed to contain concentrations of25-hydroxyvitamin D₂ and/or 25-hydroxyvitamin D₃ of 1 to 1000 μg perunit dose, for example, and are prepared in such a manner as to effectcontrolled or substantially constant release of the 25-hydroxyvitaminD₂/25-hydroxyvitamin D₃, optionally into the ileum of thegastrointestinal tract, of humans or animals over an extended period oftime. Example dosages include 1 μg to 1000 μg per unit dose, 1 μg to 600μg, 1 μg to 400 μg, 1 μg to 200 μg, 1 μg to 100 μg, 5 μg to 90 μg, 30μg, to 80 μg, 20 μg to 60 μg, 30 μg to 60 μg, 35 μg to 50 μg, 5 μg to 50μg, and 10 μg to 25 μg, for example 20 μg, 25 μg, 30 μg, 40 μg, 50 μg,60 μg, 70 μg, 80 μg, 90 μg, and 100 μg.

In one preferred class of embodiments, the controlled releaseformulation releases at least 70%, more preferably at least 80% of thevitamin D compound within the first 24 hours after dosing.

Advantageously, 25-hydroxyvitamin D₂, 25-hydroxyvitamin D₃ orcombinations thereof together with other therapeutic agents can beadministered, e.g. orally, in accordance with the above describedembodiments in dosage amounts of from 1 to 100 μg per day, for example.In one type of embodiment, the dose will be selected to provide anaverage rise in serum 25-hydroxyvitamin D₃ of about 1 to 3 ng/ml in adose interval.

In embodiments, the formulations described herein can be administered toraise and preferably also maintain blood 1,25-dihydroxyvitamin D levelsat 25 pg/mL, 30 pg/mL, or higher, e.g. 25-65 pg/mL for an extendedperiod, for example at least one month, at least three months, at leastsix months, or longer.

In one aspect, the formulations described herein can be administered topatients to lower or maintain lowered serum parathyroid hormone levels,preferably an amount that lowers PTH levels by at least 30%, oralternatively the amount needed to reduce serum levels of PTH to thetarget range for the CKD stage (e.g., for Stage 3 is 35-70 pg/mL(equivalent to 3.85-7.7 pmol/L), for Stage 4 is 70-110 pg/mL (equivalentto 7.7-12.1 pmol/L), and for Stage 5 is 150-300 pg/mL (equivalent to16.5-33.0 pmol/L) (defined in K/DOQI Guideline No. 1)).

In another aspect, the formulations according to the disclosure hereincan be administered to a patient suffering from hyperparathyroidismsecondary to chronic kidney disease (e.g., Stage 3 or 4, or Stage 3, 4or 5) to lower the serum PTH level.

The dosages described herein are contemplated for any of the therapeuticmethods described herein. It will be appreciated that the actualpreferred amount of a vitamin D compound in a specific case will varyaccording the particular compositions formulated, the mode ofapplication, and the particular situs being treated. Dosages can bedetermined using conventional considerations, e.g., by customarycomparison of the differential activity of the hormone and of a knownagent, e.g. by means of an appropriate conventional pharmacologicalprotocol.

The specific doses for each particular patient can depend on a widevariety of factors, for example, on the age, body weight, general stateof health, sex, on the diet, on the timing and mode of administration,on the rate of excretion, and on medicaments used in combination and theseverity of the particular disorder to which the therapy is applied.

Patients in need of vitamin D supplementation include healthy subjectsand subjects at risk for vitamin D insufficiency or deficiency, forexample, subjects with Stage 1, 2, 3, 4 or 5 CKD; infants, children andadults that do not drink vitamin D fortified milk (e.g. lactoseintolerant subjects, subjects with milk allergy, vegetarians who do notconsume milk, and breast fed infants); subjects with rickets; subjectswith dark skin (e.g., in the U.S., 42% of African American women between15 and 49 years of age were vitamin D deficient compared to 4% of whitewomen); the elderly (who have a reduced ability to synthesize vitamin Dand also are more likely to stay indoors); institutionalized adults (whoare likely to stay indoors, including subjects with Alzheimer's diseaseor mentally ill); subjects who cover all exposed skin (such as membersof certain religions or cultures); subjects who always use sunscreen(e.g., the application of sunscreen with a Sun Protection Factor (SPF)value of 8 reduces production of vitamin D by 95%, and higher SPF valuesmay further reduce vitamin D); subjects with fat malabsorption syndromes(including but not limited to cystic fibrosis, cholestatic liverdisease, other liver disease, gallbladder disease, pancreatic enzymedeficiency, Crohn's disease, inflammatory bowel disease, sprue or celiacdisease, or surgical removal of part or all of the stomach and/orintestines); subjects with inflammatory bowel disease; subjects withCrohn's disease; subjects who have had small bowel resections; subjectswith gum disease; subjects taking medications that increase thecatabolism of vitamin D, including phenytoin, fosphenytoin,phenobarbital, carbamazepine, and rifampin; subjects taking medicationsthat reduce absorption of vitamin D, including cholestyramine,colestipol, orlistat, mineral oil, and fat substitutes; subjects takingmedications that inhibit activation of vitamin D, includingketoconazole; subjects taking medications that decrease calciumabsorption, including corticosteroids; subjects with obesity (vitamin Ddeposited in body fat stores is less bioavailable); subjects withosteoporosis; patients who have low bone mineral density andosteoporosis; and/or postmenopausal women. According to the Institute ofMedicine's report on the Dietary Reference Intakes for vitamin D, foodconsumption data suggest that median intakes of vitamin D for bothyounger and older women are below current recommendations; data suggestthat more than 50% of younger and older women are not consumingrecommended amounts of vitamin D.

Optionally excluded from the methods of the invention described hereinare therapeutic treatment of subjects suffering from renalosteodystrophy (including osteomalacia and osteitis fibrosa cystica).

In other aspects, the compositions and methods of the invention areuseful for prophylactic or therapeutic treatment of vitamin D-responsivediseases, i.e., diseases where vitamin D, 25-hydroxyvitamin D or activevitamin D (e.g., 1,25-dihydroxyvitamin D) prevents onset or progressionof disease, or reduces signs or symptoms of disease. Such vitaminD-responsive diseases include cancer (e.g., breast, lung, skin,melanoma, colon, colorectal, rectal, prostate and bone cancer).1,25-dihydroxyvitamin D has been observed to induce cell differentiationand/or inhibit cell proliferation in vitro for a number of cells.Vitamin D-responsive diseases also include autoimmune diseases, forexample, type I diabetes, multiple sclerosis, rheumatoid arthritis,polymyositis, dermatomyositis, scleroderma, fibrosis, Grave's disease,Hashimoto's disease, acute or chronic transplant rejection, acute orchronic graft versus host disease, inflammatory bowel disease, Crohn'sdisease, systemic lupus erythematosis, Sjogren's Syndrome, eczema andpsoriasis, dermatitis, including atopic dermatitis, contact dermatitis,allergic dermatitis and/or chronic dermatitis. Vitamin D-responsivediseases also include other inflammatory diseases, for example, asthma,chronic obstructive pulmonary disease, polycystic kidney disease,polycystic ovary syndrome, pancreatitis, nephritis, hepatitis, and/orinfection. Vitamin D-responsive diseases have also been reported toinclude hypertension and cardiovascular diseases. Thus, the inventioncontemplates prophylactic or therapeutic treatment of subjects at riskof or suffering from cardiovascular diseases, for example, subjects withatherosclerosis, arteriosclerosis, coronary artery disease,cerebrovascular disease, peripheral vascular disease, myocardialinfarction, myocardial ischemia, cerebral ischemia, stroke, congestiveheart failure, cardiomyopathy, obesity or other weight disorders, lipiddisorders (e.g. hyperlipidemia, dyslipidemia including associateddiabetic dyslipidemia and mixed dyslipidemia hypoalphalipoproteinemia,hypertriglyceridemia, hypercholesterolemia, and low HDL (high densitylipoprotein)), metabolic disorders (e.g. Metabolic Syndrome, Type IIdiabetes mellitus, Type I diabetes mellitus, hyperinsulinemia, impairedglucose tolerance, insulin resistance, diabetic complication includingneuropathy, nephropathy, retinopathy, diabetic foot ulcer andcataracts), and/or thrombosis.

Diseases which can benefit from a modulation in the levels of vitamin Dcompounds, include, but are not limited to: (i) in theparathyroid—hypoparathyroidism, pseudohypo-parathyroidism, secondaryhyperparathyroidism; (ii) in the pancreas—diabetes; (iii) in thethyroid—medullary carcinoma; (iv) in the skin—psoriasis; wound healing;(v) in the lung—sarcoidosis and tuberculosis; (vi) in the kidney—chronickidney disease, hypophosphatemic VDRR, vitamin D dependent rickets;(vii) in the bone—anticonvulsant treatment, fibrogenisis imperfectaossium, osteitis fibrosa cystica, osteomalacia, osteoporosis,osteopenia, osteosclerosis, renal osteodytrophy, rickets; (viii) in theintestine—glucocorticoid antagonism, idopathic hypercalcemia,malabsorption syndrome, steatorrhea, tropical sprue; and (ix) autoimmunedisorders.

In embodiments of the invention, the disease that benefits from amodulation in the levels of vitamin D compounds are selected fromcancer, dermatological disorders (for example, psoriasis), parathyroiddisorders (for example, hyperparathyroidism and secondaryhyperparathyroidism), bone disorders (for example, osteoporosis) andautoimmune disorders.

The formulation can be prepared by procedures well within thecapabilities of the ordinary skilled artisan. For example, thecomponents of the matrix (e.g. wax and oily vehicle) can be melted, ifnecessary, to provide a flowable liquid thereby making it easier toobtain a homogeneous mixture. The active (e.g., 25-hydroxyvitamin D₂and/or 25-hydroxyvitamin D₃) is added to the liquid carrier, for exampledissolved in an alcohol such as anhydrous ethanol, and the ingredientsare mixed to provide a homogeneous mixture. In one type of embodiment,the stabilizer can be added after all matrix components (e.g., waxes andoils) are blended and prior to combination with the active. The mixturecan be cooled and stored prior to later division into unit dosage forms,such as filled gelatin capsules.

In one type of method, a portion of the oily vehicle, controlled releaseagent, and emulsifier are heated to a relatively high temperature (e.g.,65° C.) and mixed prior to adding an absorption enhancer, followed byadditional mixing until homogenous, then cooling to an intermediateelevated temperature (e.g., 50° C. to 55° C.). In a separate vessel, anantioxidant preservative and the remainder of the oily vehicle are mixedand heated to an intermediate elevated temperature (e.g., 50° C.), thencombined and mixed with the wax mixture until a homogenous solution isobtained. Next, the stabilizer is added, with mixing. Next, a solutionof the vitamin D compound(s) in alcohol is combined with the homogenouswaxy solution, mixed until a homogenous solution is obtained, preferablyfilled into capsules, and then cooled to room temperature. In anotherpreferred method, a portion of the oily vehicle, controlled releaseagent, and emulsifier are heated at a temperature of 55° C. to 60° C.and mixed prior to adding an absorption enhancer, followed by additionalmixing until homogenous. In a separate vessel, an antioxidantpreservative, the remainder of the oily vehicle, and the stabilizer aremixed and heated to a temperature of 55° C. to 60° C., then combined andmixed with the wax mixture until a homogenous solution is obtained.Next, a solution of vitamin D compound in alcohol is combined with thehomogenous waxy solution, mixed until a homogenous solution is obtained,preferably filled into capsules, and then cooled to room temperature.

The formulation preferably is placed in capsules prior to administrationto the patient in need of treatment. Such capsules may be hard or soft,and soft capsules are particularly contemplated. The formulation may befilled into gelatin capsules using standard capsule filling machinery,such as by melting the formulation and injection-filling it into softcapsule shells. Example soft capsule shells include VEGICAPS andOPTISHELL technologies (Catalent, Somerset, N.J., USA). In thealternative, the formulation can be made into a unit dosage form by anyother suitable processes, for example to yield tablets, sachets,dragees, suppositories, or the like.

In one type of embodiment, the formulation is prepared for andadministered by oral delivery. In another type of embodiment, theformulation is prepared for and administered as a suppository, e.g. arectal suppository.

The formulation and methods of use and making are contemplated toinclude embodiments including any combination of one or more of theadditional optional elements, features, and steps further describedbelow, unless stated otherwise.

Thus, in one type of embodiment, the formulation further includes apreservative, such as an antioxidant. Butylated hydroxytoluene (BHT) ispreferred.

In another type of embodiment, the vitamin D compound is administered incombination with one or more other therapeutic agents.

If the vitamin D compound is administered in combination with one ormore other therapeutic agents, the proportions of each of the compoundsin the combination being administered will be dependent on theparticular disease state being addressed. For example, one may choose toadminister 25-hydroxyvitamin D₂ and/or 25-hydroxyvitamin D₃ (e.g.,orally) with one or more calcium salts (intended as a calcium supplementor dietary phosphate binder), bisphosphonates, calcimimetics, nicotinicacid, iron, phosphate binders, cholecalciferol, ergocalciferol, activeVitamin D sterols, glycemic and hypertension control agents, variousantineoplastic agents and inhibitors of CYP24 and other cytochrome P450enzymes that can degrade vitamin D agents. In addition, one may chooseto intravenously administer 25-hydroxyvitamin D₂ and/or25-hydroxyvitamin D₃ with cholecalciferol, ergocalciferol, activeVitamin D sterols, glycemic and hypertension control agents, variousantineoplastic agents and inhibitors of CYP24 and other cytochrome P450enzymes that can degrade vitamin D agents. In practice, higher doses ofthe compounds of the present invention are used where therapeutictreatment of a disease state is the desired end, while the lower dosesare generally used for prophylactic purposes, it being understood thatthe specific dosage administered in any given case will be adjusted inaccordance with the specific compounds being administered, the diseaseto be treated, the condition of the subject and the other relevantmedical facts that may modify the activity of the drug or the responseof the subject, as is well known by those skilled in the art.

As described above, the formulation is preferably filled into gelatincapsules, but it may also be administered in neat form, or with one ormore external coating layers, such as an enteric coating. It is alsocontemplated that the formulation can be pressed into tablets, and insuch cases one or more tablet pressing excipients may be included.

In the compositions and methods described herein, preferred steps,preferred components, preferred compositional ranges thereof, andpreferred combinations of the foregoing, can be selected from thevarious specific examples provided herein. For example, a preferredformulation includes 25-hydroxyvitamin D (e.g., about 30 μg, about 60 orabout 90 μg 25-hydroxyvitamin D₃), about 2 wt % (e.g., 2.32 wt %)anhydrous ethanol, about 10 wt % (e.g., 9.75 wt %) GELUCIRE 44/14, about20 wt % (e.g., 20.00 wt. %) hard paraffin, about 23 wt % (e.g., 22.55 wt%) GMS, about 35 wt % (e.g., 35.36 wt %) liquid paraffin or mineral oil,about 10 wt % HPMC, and optionally a small amount of preservative (e.g.,0.02 wt % BHT). A variation on this formulation will include about 15%(e.g., 15.29 wt %) HPMC and about 30 wt % (e.g., 29.88 wt %) liquidparaffin or mineral oil.

Examples

The following Examples illustrate specific formulations and methods fortheir preparation. The Examples are provided for illustration and arenot intended to limit the scope of the invention.

In vitro dissolution testing in the Examples was performed using USPApparatus 2 (paddle method) as described in USP 29-NF 24, generalchapter <711> Dissolution, using the Dissolution Medium described below.In general, the method proceeds according to the following steps. Placethe stated volume of the Dissolution Medium (±1%) in the vessel of thespecified apparatus, assemble the apparatus, equilibrate the DissolutionMedium to 37±0.5°, and remove the thermometer. Place the dosage units inthe apparatus, taking care to exclude air bubbles from the surface ofthe dosage units, and immediately operate the apparatus at the specifiedrate. At each of the times stated, withdraw specimens from a zone midwaybetween the surface of the Dissolution Medium and the top of therotating blade, not less than 1 cm from the vessel wall. Replace thealiquots withdrawn for analysis with equal volumes of fresh DissolutionMedium at 37° or. Keep the vessel covered for the duration of the test,and verify the temperature of the mixture under test at suitable times.Perform the analysis using a suitable assay method, ultra performanceliquid chromatography (UPLC) in this case.

Six capsules of each formulation were tested per time point. TheDissolution Medium was 0.05 M pH 6.8 phosphate buffer/1% sodium dodecylsulfate dissolution medium at 37±0.5° C., and the apparatus was operatedat 100 rotations per minute. Samples were taken at 2, 4, 6, 8, and 12hours and the 25-hydroxyvitamin D content of each sample was determinedusing UPLC.

Example 1—In Vitro Dissolution of Unstabilized Sustained ReleaseFormulations of 25-Hydroxyvitamin D

The dissolution of a formulation made from a mixture of 90 μg of25-hydroxyvitamin D₃, 19.98 wt % hard paraffin, 37.85 wt % GMS, 9.76 wt% GELUCIRE 44/14, 2.36 wt % anhydrous ethanol, 29.88 wt % liquidparaffin, and 0.02 wt % BHT (Comparative Formulation 1) was tested. Theformulation did not comprise a cellulosic compound. The mean amount of25-hydroxyvitamin D₃ released, calculated as a mean percentage of thenominal drug loading per dosage form (mean % of label claim, % LC) atT=0 and after controlled storage of the formulation at 5° C. and ambienthumidity for up to 12 months are summarized in the table below. It wasdetermined that the samples were stored for a period of about 3 monthsat a temperature in a range of 15° C. to 30° C. and ambient humidity,prior to testing. Thus, the sample which should have represented timezero is labeled as T=0_(p) (pseudo time zero), and it should beunderstood that the nominal 1 month, 3 month, 6 month, 9 month, and 12month aged samples also experienced the approximately 3-month agingperiod just described. To provide a more accurate baseline, a freshbatch of the same type of samples was prepared and tested without anyaging; this data is labeled as T=O_(f) to indicate fresh samples. Thecoefficient of variation (% CV) is also reported. The percent changefrom the initial amount of 25-hydroxyvitamin D₃ released by T=0_(p) andT=0_(f) lots is provided in brackets and double brackets, respectively.

Dissolution Followin Storage at 5° C./Ambient Humidity 1 Month 3 Months6 months 9 months 12 months (% CV) (% CV) (% CV) (% CV) (% CV) [% change[% change [% change [% change [% change from T = 0_(p)] from T = 0_(p)]from T = 0_(p)] from T = 0_(p)] from T = 0_(p)] Time T = 0_(p) T = 0_(f)[[% change [[% change [[% change [[% change [[% change (hours) pseudofresh from T = 0_(f)]] from T = 0_(f)]] from T = 0_(f)]] from T =0_(f)]] from T = 0_(f)]] 2 6.1 22.1 8.9 15.7 9.1 15.2 12.7 (29.6) (17.2)(13.4) (27.4) (23) (47.2) (16.8) [45.9] [157.4] [49.2] [149.2] [108.2][[59.7]] [[29.0]] [[58.8]] [[31.2]] [[42.5]] 4 14.5 52.0 20.7 22.3 22.925.2 24.7 (15.7) (4.6) (19.1) (15.6) (7.1) (19.2) (13.5) [42.8] [53.8][57.9] [73.8] [70.3] [[60.2]] [[57.1]] [[56.0]] [[51.5]] [[52.5]] 6 27.677.9 35.7 33.5 34.1 36.2 34.8 (21.2) (4.6) (9.6) (2.6) (7.1) (20.5)(14.6) [29.3] [21.4] [23.6] [31.2] [26.1] [[54.2]] [[57.0]] [[56.2]][[53.5]] [[55.3]] 8 45.7 96.8 53.0 47.4 46.8 47.8 46.4 (23.6) (2.9)(9.9) (3.7) (6) (18.6) (8.9) [16.0] [3.7] [2.4] [4.6] [1.5] [[45.2]][[51.0]] [[51.7]] [[50.6]] [[52.1]] 12 89.7 112.3 100.0 78.9 76.9 74.178.8 (15.6) (1.6) (4.8) (8.1) (5.7) (17.3) (5.3) [11.5] [12.0] [14.3][17.4] [12.2] [[11.0]] [[29.7]] [[31.5]] [[34.0]] [[29.8]]

The dissolution of Comparative Formulation 1 after storage at 25° C. and60% relative humidity for 0 to 12 months was tested. The results aresummarized in the table below.

Dissolution Following Storage at 25° C./60% Relative Humidity 1 Month 3Months 6 months 9 months 12 months (% CV) (% CV) (% CV) (% CV) (% CV) [%change [% change [% change [% change [% change from T = 0_(p)] from T =0_(p)] from T = 0_(p)] from T = 0_(p)] from T = 0_(p)] [[% change [[%change [[% change [[% change [[% change Dissolution T = 0_(p) T = 0_(f)from from from from from Time (hours) pseudo fresh T = 0_(f)]] T =0_(f)]] T = 0_(f)]] T = 0_(f)]] T = 0_(f)]] 2 6.1 22.1 7.6 10.8 8.5 10.813.5 (29.6) (17.2) (7.7) (15.7) (19.2) (15.8) (24.7) [24.6] [77.0][39.3] [77.0] [121.3] [[65.6]] [[51.1]] [[61.5]] [[51.1]] [[38.9]] 414.5 52.0 18.7 22.8 17.9 21.4 24.5 (15.7) (4.6) (18.6) (23.7) (11.9)(5.5) (17.4) [29.0] [57.2] [23.4] [47.6] [69.0] [[64.0]] [[56.2]][[65.6]] [[58.8]] [[52.9]] 6 27.6 77.9 27.1 30.7 23.8 27.0 30.0 (21.2)(4.6) (22.7) (29.6) (11.6) (7.3) (15.7) [1.8] [11.2] [13.8] [2.2] [8.7][[65.2]] [[60.6]] [[69.4]] [[65.3]] [[61.5]] 8 45.7 96.8 37.1 40.6 28.532.3 35.6 (23.6) (2.9) (18.1) (29.9) (13.2) (6.4) (14.7) [18/8] [11.2][37.6] [29.3] [22.1] [[61.7]] [[58.1]] [[70.6]] [[66.6]] [[63.2]] 1289.7 112.3 61.6 53.0 38.5 38.9 44.2 (15.6) (1.6) (16.6) (32.2) (12.2)(6) (12.2) [31.3] [40.9] [57.1] [56.6] [50.7] [[45.1]] [[52.8]] [[65.7]][[65.4]] [[60.6]]

The dissolution of Comparative Formulation 1 after storage at 40° C. and75% relative humidity for 0, 1, 3, and 6 months was tested. The resultsare summarized in the table below.

Dissolution Following Storage at 40° C./75% Relative Humidity 1 Month 3Months 6 Months (% CV) (% CV) (% CV) [% change [% change [% change fromfrom from Dissolution T = 0_(p)] T = 0_(p)] T = 0_(p)] Time T = 0_(p) T= 0_(f) [[% change [[% change [[% change (hours) pseudo fresh from T =0_(f)]] from T = 0_(f)]] from T = 0_(f)]] 2 6.1 22.1 11.7 16.9 1.8(29.6) (17.2) (50)   (36.3) (75)   [91.8]  [177.0] [70.5]  [[47.1]][[23.5]] [[91.9]] 4 14.5 52.0 52.0 59.8 36.2 (15.7) (4.6)  (45.1) (31.3)(21.8) [258.6] [312.4] [149.7] [[0]]   [[15.0]] [[30.4]] 6 27.6 77.987.0 97.8 76.7 (21.2) (4.6)  (21.5) (24)   (12.3) [215.2] [254.3][177.9] [[11.7]] [[25.5]] [[1.5]]  8 45.7 96.8 107.3 110.9 101.2 (23.6)(2.9)  (8.1)  (13)   (6.1)  [134.8] [142.7] [121.4] [[10.8]] [[14.6]][[4.5]]  12 89.7 112.3 118.7 115.1 112.6 (15.6) (1.6)  (1.7)  (3.7) (2.1)  [32.3]  [28.3]  [25.5]  [[5.7]]  [[2.5]]  [[0.3]] Without intending to be bound by any particular theory, the increase inthe extent of dissolution following storage at 40° C. compared to thepseudo T=0 values is believed to be due to a combination of the agingeffect described above on the pseudo T=0 samples tested, and atemperature-dependent phase change in the formulation when stored at 40°C.

Aged product according to Comparative Formulation 1 was heat cured andthen subject to dissolution testing. Curing consists of applying a heattreatment and has been shown to stabilize pharmaceutical formulations(see, e.g., U.S. Pat. No. 6,645,527), Comparative Formulation 1 (agedsamples) was heated at 40° C. for 72 hours for curing, and then wasstored at room temperature for 8 weeks. Release of 25-hydroxyvitamin D₃from the cured formulation was tested after storage for 0, 2, 4, and 8weeks at room temperature. The results are summarized in the tablebelow.

Dissolution Following Curing at 40° C. for 72 Hours Dissolution 2 weeks4 weeks 8 weeks Time % LC % LC % LC (hours) T = 0 [% change] [% change][% change] 2 17.4 12.4 11.4 8.2 [28.7] [34.5] [52.9] 4 53.3 46.4 40.126.6 [12.9] [24.8] [50.1] 6 86.2 76.2 69.0 44.8 [11.6] [20.0] [48.0] 8103.8 102.0 95.8 66.6 [1.7]  [7.7]  [35.8] 12 115.7 110.5 119.8 103.3[4.5]  [3.5]  [10.7]

Example 2—In Vitro Dissolution of Stabilized Controlled ReleaseFormulations of 25-Hydroxyvitamin D

The dissolution of a sustained release formulation comprising 90 μg of25-hydroxyvitamin D₃, 19.88 wt % hard paraffin, 15.29 wt % hydroxypropylmethylcellulose, 22.55 wt % GMS, 9.76 wt % GELUCIRE 44/14, 2.36 wt %anhydrous ethanol, 29.88 wt % liquid paraffin, and 0.02 wt % BHT(Example Formulation A) was tested after 0 to 11 weeks of storage atroom temperature. The results are summarized in the table below.

Dissolution Following Storage at Room Temperature/Ambient HumidityDissolution 3 weeks 11 weeks Time % LC % LC (hours) T = 0 [% change] [%change] 2 15.45 14.20 12.08 [8.1%]  [21.8%] 4 36.3 38.80 37.13 [6.9%] [2.3%]  6 56.9 62.70 59.51 [10.2%] [4.6%]  8 71.1 71.90 69.76 [1.1%] [1.9%]  12 89.4 91.20 89.90 [2.0%]  [0.6%] 

The dissolution of a sustained release formulation comprising 90 μg of25-hydroxyvitamin D₃, 19.88 wt % hard paraffin, 10.00 wt % hydroxylpropyl methylcellulose, 22.55 wt % GMS, 9.76 wt % GELUCIRE 44/14, 2.36wt % anhydrous ethanol, 35.17 wt % liquid paraffin, and 0.02 wt % BHT(Example Formulation B) was tested after 0 to 26 weeks of storage atroom temperature. The results are summarized in the table below.

Dissolution Following Storage at Room Temperature/Ambient Humidity 6 1326 Dis- weeks weeks weeks solution % LC % LC % LC Time [% [% [% (hours)T = 0 change] change] change] 2 30.15 25.40 25.20 21.10 [15.8%] [16.4%][30.0%] 4 58.55 51.90 51.80 45.20 [11.4%] [11.5%] [22.8%] 6 72.1 74.4073.00 67.63 [3.2%]  [1.2%]  [6.2%]  8 80.55 84.30 84.50 77.30 [4.7%] [4.9%]  [4.0%]  12 91.8 94.10 94.40 91.16 [2.5%]  [2.8%]  [0.7%] Example Formulation B demonstrated a substantially stable dissolutionprofile following storage for at least 26 weeks at room temperature.

The stability of stabilized formulations comprising 30 μg (ExampleFormulation C), 60 μg (Example Formulation D), or 90 μg (ExampleFormulation E) of 25-hydroxyvitamin D₃ was tested using storageconditions of 25° C. and 60% relative humidity and 40° C. and 75%relative humidity. The compositions of Example Formulations C to E aresummarized in the table below:

Component Amount 25-hydroxyvitamin D₃ 30 μg, 60 μg, or 90 μg ParaffinWax 20.00 wt % Mineral Oil 35.36 wt % Hydroxy propyl 10.00 wt %Methylcellulose K100M CR (METHOCEL) Glycerol monostearate 22.55 wt %Lauroyl macrogolglycerides and  9.75 wt % polyoxylglycerides (GELUCIRE44/14) Anhydrous Alcohol  2.32 wt % BHT  0.02 wt % Soft Capsule Shell(VEGICAPS)The formulations exhibited substantially stable dissolution profilesfollowing storage at 25° C. and 60% relative humidity for at least 24months (FIG. 1). The dissolution results (% LC and % CV) are summarizedin the table below.

Dissolution Following Storage at 25° C./60% Relative Humidity T = 0 1month 3 mos. 6 mos. 9 mos. 12 mos. 18 mos. 24 mos. Time % LC % LC % LC %LC % LC % LC % LC % LC (hours) (% CV) (% CV) (% CV) (% CV) (% CV) (% CV)(% CV) (% CV) 30 μg 25-hydroxyvitamin D₃ (Example Formulation C) 2 10.112.9 14.0  9.6 10.5 13.5 10.6  7.0 (16.1) (25.2) (48.5) (38.3) (18.2)(35.8) (26.5) (47.8) 4 43.6 48.8 45.9 36.8 37.2 50.2 39.3 39.6 (12.7)(10.9) (29.9) (25)   (11.1) (14.8) (22.9) (32.3) 6 83.1 73.4 72.4 71.466.7 70.6 70.1 69.3  (5.7) (9.2) (7.5)  (6.8) (10.6) (11.5)  (7.2)(12.6) 8 96.4 89.6 88.4 88.4 85.2 85.4 84   85.7  (6.5) (4.4) (4.7) (5.7)  (4.9) (9)   (5.1)  (8.2) 12 115.8  104.2 105.6 106.1  101.5 100.8  100.2  103    (4.5) (1.9) (1.4)  (1.6)  (1.5)  (3.9)  (4.5) (2.6) 60 μg 25-hydroxyvitamin D₃ (Example Formulation D) 2 17.5 16.316.1 17.7 11.2 14.3 16.1 12.4 (20)   (27.4) (25.3) (34.7) (20.2) (29.1)(25.8) (52.3) 4 53.6 55.1 53.8 55.3 43.7 51   52.9 41   (19.6) (18.8)(12.8) (17.9) (14)   (22.5) (16.7) (36.6) 6 83.9 78.7 79.9 78.4 72.275   72   64.7  (7.5) (8.3) (6.6)  (4.7)  (7.1) (13)    (9.9) (29.6) 899.2 94.3 97.2 92.5 87   88.9 84.9 81.3  (3.9) (5.2) (4.8)  (3.3)  (5.5) (6.3)  (12.3)* (21.7) 12 104.8  108.7 111.9 104.5  103.1  104.7  99.8101.7   (3.3) (1.8) (1.1)  (0.5)  (0.8)  (1.4)  (4.6)* (8)  90 μg25-hydroxyvitamin D₃ (Example Formulation E) 2 14.9 14.9 13.3 13.5 14.315    9.5  8.5 (19.9) (8.6) (41.2) (23.4) (30.4) (35.1) (37.1) (53.2) 449.9 46.9 52.4 49.7 45.9 51.8 36.8 34.4 (16.4) (10.2) (18.5) (16.4)(25.6) (14)   (34.5) (24.9) 6 89.4 71.4 81.1 74.2 71.6 77   64.4 64.5 (7.2) (4.1) (5.6) (11.1) (17.6)  (4.9) (10.3) (15.1) 8 101.7  84.9 96.190.8 89   91.6 77.4 83.6  (2.5) (2.8) (2.3)  (6.4)  (9.4)  (3.3) (15.7)(11.2) 12 103   99.3 110 104.4  100.5  104.6  96.8 102.8   (2.1) (2.4)(1.3)  (1.1)  (2.4)  (0.3) (5)   (3.4) *4 replicates instead of 6

The percent change between the amount of 25-hydroxyvitamin D₃ releasedfollowing aging compared to the initial amount released is summarized inthe table below.

Time 1 month 3 mos. 6 mos. 9 mos. 12 mos. 18 mos. 24 mos. (h) % change %change % change % change % change % change % change 30 μg25-hydroxyvitamin D₃ (Example Formulation C) 2 27.7% 38.6% 5.0% 4.0%33.7% 5.0% 30.7% 4 11.9% 5.3% 15.6% 14.7% 15.1% 9.9% 9.2% 6 11.7% 12.9%14.1% 19.7% 15.0% 15.6% 16.6% 8 7.1% 8.3% 8.3% 11.6% 11.4% 12.9% 11.1%12 10.0% 8.8% 8.4% 12.3% 13.0% 13.5% 11.1% 60 μg 25-hydroxyvitamin D₃(Example Formulation D) 2 6.9% 8.0% 1.1% 36.0% 18.3% 8.0% 29.1% 4 2.8%0.4% 3.2% 18.5% 4.9% 1.3% 23.5% 6 6.2% 4.8% 6.6% 13.9% 10.6% 14.2% 22.9%8 4.9% 2.0% 6.8% 12.3% 10.4% 14.4% 18.0% 12 3.7% 6.8% 0.3% 1.6% 0.1%4.8% 3.0% 90 μg 25-hydroxyvitamin D₃ (Example Formulation E) 2 0.0%10.7% 9.4% 4.0% 0.7% 36.2% 43.0% 4 6.0% 5.0% 0.4% 8.0% 3.8% 26.3% 31.1%6 20.1% 9.3% 17.0% 19.9% 13.9% 28.0% 27.9% 8 16.5% 5.5% 10.7% 12.5% 9.9%23.9% 17.8% 12 3.6% 6.8% 1.4% 2.4% 1.6% 6.0% 0.2%

Example Formulations C to E also exhibited substantially stabledissolution profiles following storage at 40° C. and 75% RH for at least6 months (FIG. 2). The dissolution results are summarized in the tablebelow.

Dissolution Following Storage at 40° C./75% Relative Humidity Initial 1month 3 months 6 months Time % LC % LC % LC % LC (hours) (% CV) (% CV)(% CV) (% CV) 30 μg 25-hydroxyvitamin D₃ (Example Formulation C) 2 10.111.3 13.5 11.1 (16.1) (54.3) (53.8) (14.8) 4 43.6 46 44.4 40.9 (12.7)(27.1) (23.9) (13.9) 6 83.1 72.7 62.8 68.7 (5.7)  (9.7)  (20.4) (7.3)  896.4 88.7 76.5 82.5 (6.5)  (5)   (12.5) (4.1)  12 115.8 103.1 93.4 96.4(4.5)  (0.9)  (7.6)  (2.8)  60 μg 25-hydroxyvitamin D₃ (ExampleFormulation D) 2 17.5 12.9 15.5 13.9 (20)   (40.7) (37)   (40)   4 53.650.7 54.4 49.8 (19.6) (18.5) (12)   (16.3) 6 83.9 75.8 78.3 74.7 (7.5) (9.7)  (10.6) (8.1)  8 99.2 91.2 91.6 88.5 (3.9)  (7.9)  (9.1)  (6.5) 12 104.8 100.7 104.1 101.5 (3.3)  (5.1)  (6.5)  (2.3)  90 μg25-hydroxyvitamin D₃ (Example Formulation E) 2 14.9 18.5 10.2 8.3 (19.9)(32.5) (44.6) (34.7) 4 49.9 50.7 47 44.8 (16.4) (12.8) (14.6) (10.6) 689.4 74.7 72.9 73 (7.2)  (8)   (5.2)  (3.4)  8 101.7 90.5 86.6 87.9(2.5)  (5)   (4.8)  (3)   12 103 100.1 102.5 101.0 (2.1)  (1.4)  (1.6) (1.9) 

The percent change between the amount of 25-hydroxyvitamin D₃ releasedfollowing exposure to storage conditions compared to the initial amountreleased is summarized in the table below.

Time 1 month 3 months 6 months (hours) % change % change % change 30 μg25-hydroxyvitamin D₃ (Example Formulation C) 2 11.9% 33.7%  9.9% 4  5.5% 1.8%  6.2% 6 12.5% 24.4% 17.3% 8  8.0% 20.6% 14.4% 12 11.0% 19.3% 16.8%60 μg 25-hydroxyvitamin D₃ (Example Formulation D) 2 26.3% 11.4% 20.6% 4 5.4%  1.5%  7.1% 6  9.7%  6.7% 11.0% 8  8.1%  7.7% 10.8% 12  3.9%  0.7% 3.1% 90 μg 25-hydroxyvitamin D₃ (Example Formulation E) 2 24.2% 31.5%44.3% 4  1.6%  5.8% 10.2% 6 16.4% 18.5% 18.3% 8 11.0% 14.8% 13.6% 12 2.8%  0.5%  1.9%

The stability of Comparative Formulation 1, which does not contain acellulosic compound, and Example Formulation E comprising hydroxylpropyl methylcellulose was evaluated following storage for 12 months at25° C. and 60% relative humidity (FIG. 3). The dissolution results aresummarized in the table below.

Dissolution Following Storage at 25° C./60% Relative HumidityComparative Example Dissolution Formulation 1 Formulation E Time Initial12 months Initial 12 months (hours) % LC % LC % LC % LC 2 22.1 13.5 14.915.0 4 52.2 24.5 49.9 51.8 6 77.9 30.0 89.4 77.0 8 96.8 35.6 101.7 91.612 112.3 44.2 103.0 104.6

The percent change between the amount of 25-hydroxyvitamin D₃ releasedfollowing exposure to storage conditions compared to the initial amountreleased is summarized in the table below.

Comparative Example Dissolution Formulation 1 Formulation E Time %change % change (hours) from initial from initial 2 38.9% 0.7% 4 53.1%3.8% 6 61.5% 13.9%  8 63.2% 9.9% 12 60.6% 1.6%

Example 3: In Vivo Results for Unstabilized and Stabilized ControlledRelease Formulations

In vivo studies were conducted to evaluate the clinical pharmacokineticsof unstabilized and stabilized controlled release formulations of25-hydroxyvitamin D₃ in human subjects. In Study A, 28 subjects withstage 3 or stage 4 CKD, secondary hyperparathyroidism (stage 3: 70-1000pg/mL iPTH; stage 4: 110-1000 pg/mL iPTH), and vitamin D insufficiency(serum total baseline 25-hydroxyvitamin D of 15 ng/mL to 29 ng/mL)received a single oral dose of a controlled release capsule comprising450 μg or 900 μg of 25-hydroxyvitamin D₃, 20.00 wt % hard paraffin,37.85 wt % GMS, 9.75 wt % GELUCIRE 44/14, 2.32 wt % anhydrous ethanol,30.06 wt % mineral oil, and 0.02 wt % BHT (Comparative Formulation 3) ora single intravenous dose of 448 μg 25-hydroxyvitamin D₃ in an ethanolsolution. None of the formulations comprised a cellulosic compound.

The serum concentration of 25-hydroxyvitamin D₃ increased graduallyfollowing the administration of an oral dose. The increase in25-hydroxyvitamin D₃ was dose proportional and reached an approximatemean maximum observed serum concentration (Cmax) of 32 ng/mL followingthe administration of the 900 μg capsule. The time at which Cmaxoccurred (Tmax) was approximately 13 hours post-dose. In contrast,concentrations of 25-hydroxyvitamin D₃ increased rapidly following theadministration of the i.v. dose. Peak serum levels were achievedimmediately following administration of the i.v. dose (Tmax=0.5 hours)and reached an approximate mean Cmax of 134 ng/mL. The bioavailabilityof the oral doses was approximately 6 to 11%. The terminal half-life(t_(1/2)) of 25-hydroxyvitamin D₃ following the administration of theoral dose was approximately 12 to 22 days. No adverse effects on serumcalcium or phosphorous, or urine calcium were observed in any treatmentgroup.

The mean serum total 1,25-dihydroxyvitamin D rose rapidly following theadministration of the i.v. injection, increasing from pre-treatmentbaseline by approximately 13 pg/mL by 6 hours post-dose. In contrast,mean serum total 1,25-dihydroxyvitamin D increased dose proportionallyand gradually by approximately 7 pg/mL by 48 hours post-dose followingthe administration of the 900 μg capsule.

Serum iPTH showed no meaningful change over the first 96 hours after thei.v. dose was administered. In contrast, serum PTH declined graduallyfollowing dosing, reaching a maximum suppression of approximately 20%from pretreatment baseline for subjects who received the 900 μg capsule.The observed pharmacokinetic parameters for all the treatment groups aresummarized in the table below.

450 μg po (N = 9) 900 μg po (N = 9) 448 μg iv (N = 9) n n n Mean (SD)Mean (SD) Mean (SD) Parameter Median Range Median Range Median RangePK/PD Population Observed 25-hydroxyvitamin D₃ AUC_(0-42 days) 913353.12 (10606.47) 9 21563.95 (9165.53) 9 43463.51 (12589.57) (ng*h/mL)12196.45 17940.11 47115.66 888.39, 9292.88, 23340.59, 32885.95 38631.9163006.78 AUC_(0-last) 9 13353.12 (10606.47) 9 21563.95 (9165.53) 943463.51 (12589.57) (ng*h/mL) 12196.45 17940.11 47115.66 888.39,9292.88, 23340.59, 32885.95 38631.91 63006.78 AUC_(0-inf) 9 81511.71(103037.08) 9 122901.73 (114168.13) 9 137955.58 (66746.71) (ng*h/mL)54967.57 79902.04 123580.87 4927.95, 25729.84, 39282.49, 333366.90378935.59 243322.76 C_(max) 9 25.18 (10.134) 9 31.54 (15.765) 9 133.99(19.311) (ng/mL) 20.52 30.12 133.68 15.35, 12.21, 91.71, 42.24 67.01160.91 C_(last) 9 18.11 (7.846) 9 19.08 (7.611) 9 35.07 (12.330) (ng/mL)15.84 21.37 36.91 10.30, 7.30, 12.68, 29.80 27.93 53.39 t_(max) (h) 913.11 (9.597) 9 13.56 (9.989) 9 0.49 (0.638) 10.00 10.00 0.25 6.00,2.00, 0.083, 36.00 30.00 2.00 λ₂ (h⁻¹) 9 0.0015 (0.0028) 9 0.0003(0.0002) 9 0.0005 (0.0002) 0.0003 0.0004 0.0004 0.0001, 0.0001, 0.0002,0.0087 0.0005 0.0008 R² 9 0.89 (0.130) 9 0.90 (0.169) 9 0.91 (0.090)0.96 0.99 0.93 0.662, 0.523, 0.730, 1.000 0.998 1.000 t_(1/2) (h) 92477.72 (2581.24) 9 3228.63 (2734.74) 9 1775.86 (779.13) 2483.61 1937.321694.69 79.24, 100.13, 871.46, 8615.11 9646.71 3297.78 V_(d) 9 49.42(18.30) 9 45.06 (19.38) 9 20.35 (7.42) (L/ng) 50.93 40.080 19.550 23.20,20.77, 13.04, 72.76 87.51 32.68 CL 9 0.0499 (0.0155) 9 0.0155 (0.0119) 90.0095 (0.0065) (L/ng*h) 0.0182 0.0125 0.0081 0.0030, 0.0026, 0.0041,0.2029 0.0389 0.0255 F NA 0.306 (NA) NA 0.247 (NA) NA 1.000 (NA) NA NANA NA NA NA

The baseline-adjusted pharmacokinetic parameters for all the treatmentgroups are summarized in the table below.

450 μg po (N = 9) 900 μg po (N = 9) 448 μg iv (N = 9) n Mean (SD) n Mean(SD) n Mean (SD) Parameter Median Range Median Range Median RangeAUC_(0-42 days) 9 1394.89 (1911.41) 8 4525.43 (3123.29) 9 19609.07(5319.01) (ng*h/mL) 605.42 4801.54 18764.48 48.87, 148.07, 11066.02,4956.68 8843.74 28611.05 AUC_(0-last) 9 1257.30 (2047.74) 8 4274.27(3488.40) 9 19609.07 (5319.01) (ng*h/mL) 48.87 4801.54 18764.48 −325.43,−877.99, 11066.02, 4956.68 8843.74 28611.05 AUC_(0-inf) 6 3318.90(4606.89) 8 6791.49 (5224.54) 9 34543.75 (22103.97) (ng*h/mL) 547.256872.86 26962.25 50.78, 285.04, 16868.97, 9878.03 14979.17 89126.53C_(max) 9 6.90 (4.266) 8 14.17 (9.884) 9 110.33 (14.536) (ng/mL) 5.7012.30 111.08 2.93, 2.55, 76.63, 14.87 35.59 127.40 C_(last) 9 2.36(2.257) 8 2.64 (1.784) 9 11.40 (5.648) (ng/mL) 2.05 3.01 9.01 0.16,0.42, 5.84, 5.49 5.40 22.86 t_(max) (h) 9 13.11 (9.597) 8 15.00 (9.621)9 0.49 (0.638) 10.00 10.00 0.25 6.00, 8.00, 0.083, 36.00 30.00 2.00 λ₂(h⁻¹) 6 0.018 (0.359) 7 0.0020 (0.0013) 9 0.0011 (0.0005) 0.0042 0.00160.0010 0.0008, 0.0008, 0.0004, 0.0910 0.0037 0.0021 R² 6 0.92 (0.101) 60.99 (0.020) 9 0.92 (0.086) 0.96 0.99 0.95 0.743, 0.948, 0.720, 0.9991.000 0.999 t_(1/2) (h) 6 307.86 (336.42) 6 522.96 (320.80) 9 745.86(437.65) 165.72 530.00 663.43 7.61, 189.84, 337.37, 914.23 879.311834.71 V_(d) 6 340.42 (269.11) 6 82.92 (29.70) 9 32.94 (11.00) (L/ng)249.77 77.11 29.70 80.51, 39.96, 24.28, 771.79 127.06 58.95 CL 6 4.588(7.525) 6 0.141 (0.080) 9 0.036 (0.014) (L/ng*h) 2.119 0.1200 0.0370.101, 0.0668, 0.011, 19.692 0.2860 0.059 F NA 0.064 (NA) NA 0.109 (NA)NA 1.000 (NA) NA NA NA NA NA NA

In Study B, 20 healthy subjects with mean baseline serum25-hydroxyvitamin D of about 24 ng/mL (range 11 ng/mL to 45 ng/mL)received a single oral dose of a stabilized controlled release capsulecomprising 900 μg of 25-hydroxyvitamin D₃, 20.00 wt % hard paraffin,10.00 wt % HPMC, 22.55 wt % GMS, 9.75 wt % GELUCIRE 44/14, 2.32 wt %anhydrous ethanol, 35.36 wt % mineral oil, and 0.02 wt % BHT (ExampleFormulation F) or a single intravenous dose of 448 μg 25-hydroxyvitaminD₃ in an ethanol solution.

The gradual increase in 25-hydroxyvitamin D₃ levels was demonstrated bythe prolonged Tmax following administration of the stabilized oralformulation compared to the i.v. dose. The pharmacokinetic profilefollowing administration of the stabilized oral formulation demonstrateda gradual increase in 25-hydroxyvitamin D₃ concentrations, with a meanTmax of 28 hours, while avoiding rapid increases in blood levels in themajority of subjects. Administration of the i.v. dose resulted in arapid increase in 25-hydroxyvitamin D₃ concentrations in all subjects.The avoidance of the rapid increase in 25-hydroxyvitamin D₃ levels washighlighted by the marked difference in the observed C_(max) between thetreatment groups. The Cmax following the oral dose was 58 ng/mL,compared to a Cmax of 153 ng/mL following the i.v. dose.

The exposure to 25-hydroxyvitamin D₃ following administration of thecontrolled release capsule was approximately two-fold lower thanfollowing the i.v. dose despite the oral dose being approximatelytwo-fold higher, resulting in a bioavailability of approximately 25%.The t_(1/2), clearance (CL) and volume of distribution (Vd) appeared tobe similar between treatment groups. The values for t_(1/2) and CL wereconsistent with the reported prolonged elimination of 25-hydroxyvitaminD₃. In addition, the Vd values suggested that 25-hydroxyvitamin D₃ wasmaintained in systemic circulation, likely highly bound to the DBP. Theobserved pharmacokinetic parameters for all the treatment groups aresummarized in the table below.

900 μg CTAP101 448 μg CTAP101 Capsules Injection Parameter (N = 10) (N =10) AUC_(0-t) (ng · h/mL) Mean (SD) 21545.20 (7054.02) 25274.44(7206.93) Median 19904.00 25810.85 Minimum, 10176.40, 8434.02, Maximum35885.02 35382.55 AUC_(0-inf) (ng · h/mL) Mean (SD) 77945.13 (55896.15)55234.52 (27268.64) Median 58974.22 51247.29 Minimum, 19504.43,22979.18, Maximum 194796.33 119865.38 C_(max) (ng/mL) Mean (SD) 57.657(39.3810) 153.029 (20.8620) Median 37.925 152.890 Minimum, 24.43,125.94, Maximum 146.86 185.33 t_(max) (h) Mean (SD) 28.100 (27.4000)0.272 (0.2910) Median 21.000 0.167 Minimum, 4.00, 0.05, Maximum 96.001.00 t_(1/2) (h) Mean (SD) 1389.40 (1144.48) 660.23 (415.82) Median1042.60 607.65 Minimum, 557.32, 238.46, Maximum 4171.67 1733.68 λ₂ (h⁻¹)Mean (SD) 0.00072 (0.00036) 0.00136 (0.00071) Median 0.00067 0.00115Minimum, 0.00017, 0.00040, Maximum 0.00124 0.00291 CL (L/h) Mean (SD)0.0064 (0.0027) 0.0098 (0.0046) Median 0.0066 0.0087 Minimum, 0.0018,0.0037, Maximum 0.0098 0.0195 Vd (L) Mean (SD) 9.27 (0.97) 7.50 (1.19)Median 9.39 7.24 Minimum, 7.93, 6.02, Maximum 10.49 9.77 R² Mean (SD)0.80 (0.24) 0.92 (0.075) Median 0.88 0.94 Minimum, 0.25, 0.74, Maximum0.99 1.0 F* Mean 0.42 (0.14) N/A Median 0.39 N/A Minimum, 0.20, N/AMaximum 0.71 *average of individual subjects' bioavailabilitiesAbbreviations: N/A, not applicable

The baseline-adjusted pharmacokinetic parameters for all the treatmentgroups are summarized in the table below.

900 μg CTAP101 448 μg CTAP101 Capsules Injection Parameter (N = 10) (N =40) AUC_(0-t) (ng · h/mL) Mean (SD) 6891.81 (6678.97) 13583.95 (3908.42)Median 4360.23 14853.46 Minimum, 1017.88, 5302.50, Maximum 20340.6817194.59 AUC_(0-inf) (ng · h/mL) Mean (SD) 9418.00 (9410.58) 17735.09(5249.38) Median 5420.04 18229.25 Minimum, 1179.07, 9820.16, Maximum28031.64 25534.45 C_(max) (ng/mL) Mean (SD) 35.867 (39.3886) 133.653(20.7925) Median 14.910 133.785 Minimum, 6.50, 103.88, Maximum 120.52166.34 t_(max) (h) Mean (SD) 28.100 (27.4001) 0.272 (0.2914) Median21.000 0.167 Minimum 4.00, 0.05, Maximum 96.00 1.00 t_(1/2) (h) Mean(SD) 270.61 (215.00) 264.08 (82.23) Median 194.00 269.57 Minimum,107.90, 132.46, Maximum 832.70 382.99 λ₂ (h⁻¹) Mean (SD) 0.00362(0.00182) 0.00292 (0.00112) Median 0.00363 0.00257 Minimum, 0.00083,0.00181, Maximum 0.00624 0.00523 CL (L/h) Mean (SD) 0.027 (0.0063) 0.028(0.0093) Median 0.028 0.025 Minimum, 0.012, 0.018, Maximum 0.033 0.046Vd (L) Mean (SD) 8.78 (3.08) 9.74 (2.02) Median 8.08 9.47 Minimum, 5.06,6.54, Maximum 14.17 13.27 R² Mean (SD) 0.83 (0.22) 0.92 (0.072) Median0.88 0.94 Minimum, 0.25, 0.74, Maximum 1.0 1.0 F* Mean (SD) 0.25 (0.24)N/A Median 0.16 N/A Minimum, 0.037, N/A Maximum 0.75 *average ofindividual subjects' bioavailabilities Abbreviations: N/A, notapplicable

The study demonstrated that the stabilized controlled releaseformulation modified the rate of absorption of 25-hydroxyvitamin D₃,yielding a more gradual increase in serum 25-hydroxyvitamin D₃ levelswhile maintaining the distribution and elimination characteristics. Thestabilized formulation demonstrated improved pharmacokinetic parameters,such as increased Tmax, AUC, and bioavailability, compared to the samedose of the unstabilized formulation in Study A.

In Study C, 78 subjects with stage 3 CKD (eGFR 25-70 mL/min/1.73 m²),SHPT (>70 pg/mL plasma iPTH) and vitamin D insufficiency (serum totalbaseline 25-hydroxyvitamin D of 10 ng/mL to 29 ng/mL) received dailyoral doses of stabilized controlled release formulations comprising 30μg, 60 μg, or 90 μg of 25-hydroxyvitamin D₃, 20.00 wt % hard paraffin,10.00 wt % HPMC, 22.55 wt % GMS, 9.75 wt % GELUCIRE 44/14, 2.32 wt %anhydrous ethanol, 35.36 wt % mineral oil, and 0.02 wt % BHT (ExampleFormulations C, D, and E from Example 2) or placebo for 6 weeks.

The mean baseline serum 25-hydroxyvitamin D₃ concentrations werecomparable across treatment groups and ranged from approximately 16 to20 ng/mL. Following treatment with 25-hydroxyvitamin D₃, mean levels ofserum 25-hydroxyvitamin D₃ increased gradually and in adose-proportional manner following repeated daily administration of25-hydroxyvitamin D₃ and began to approach steady-state by 6 weeks (FIG.4). The mean baseline adjusted Cmax values were approximately 28, 60 and86 ng/mL for the groups administered 30 μg, 60 μg and 90 μg of25-hydroxyvitamin D₃, respectively. Mean exposures to 25-hydroxyvitaminD₃, assessed as background adjusted AUC_(0-6 Weeks), were doseproportional across dose groups. Following the last dose, mean serum25-hydroxyvitamin D₃ levels declined slowly, but by the end of the studyremained above baseline for all groups. Mean t_(1/2) was determined tobe approximately between 25 to 50 days. The baseline-adjustedpharmacokinetic parameters for 25-hydroxyvitamin D₃ are summarized inthe table below.

Placebo 30 μg 60 μg 90 μg Baseline (ng/mL) Mean (SD) 16.4 (8.2) 16.2(7.3) 19.8 (8.7) 18.4 (9.8) Median 12.9 17.2 21.2 16.7 Minimum, 4.4,5.0, 5.8, 6.7, Maximum 30.4 25.8 32.5 38.9 C_(max) (ng/mL) Mean (SD) 4.1(3.5) 27.8 (8.2) 60.3 (19.0) 85.7 (26.9) Median 3.1 28.1 60.8 76.0Minimum, 0.6, 10.8, 30.3, 55.4, Maximum 13.8 43.4 89.5 146.4AUC_(0-6 Weeks) (ng · d/mL) Mean (SD) 45.9 (60.0) 709.2 (246.3) 1531.4(374.8) 2134.3 (584.3) Median 32.1 6843.0 1573.0 1963.8 Minimum, −60.1,307.8, 712.7, 1377.5, Maximum 222.3 1249.0 2221.8 3207.3 t_(max) (d)Mean (SD) NA 37.8 (10.4) 41.1 (5.2) 42.6 (5.3) Median NA 42.50 43.0 43.0Minimum, NA 8.0, 29.0, 35.0, Maximum 44.0 45.0 57.0 t_(1/2) (d) Mean(SD) NA 25.8 (16.3) 33.1 (9.3) 50.1 (51.0) Median NA 24.1 31.6 37.7Minimum, NA 5.2, 17.4, 23.2, Maximum 52.6 52.3 224.0

Mean baseline serum 1,25-dihydroxyvitamin D concentrations werecomparable across treatment groups and increased gradually, similar tothe effect on serum 25-hydroxyvitamin D₃ concentrations. Mean±SDbaseline-adjusted Cmax values were higher in the 60 μg and 90 μg groups(18.4±6.24 and 19.9±14.30 ng/mL, respectively) compared to the placeboand 30 μg groups (5.7±6.35 and 6.4±7.66 ng/mL, respectively). Meanexposures to 1,25-dihydroxyvitamin D, assessed as baseline-adjustedAUC_(0-6 weeks), were dose-proportional across the 25-hydroxyvitamin D₃dose groups. The baseline-adjusted pharmacokinetic parameters for1,25-dihydroxyvitamin D are summarized in the table below.

Placebo 30 μg 60 μg 90 μg N = 23 N = 12 N = 16 N = 14 Baseline (pg/mL)Mean (SD) 20.8 (10.11) 18.3 (7.53) 20.6 (7.62) 20.6 (7.29) Median 17.017.0 18.0 21.0 Minimum, 7.0, 5.1, 8.2, 9.3, Maximum 41.4 30.7 33.6 34.5C_(max) (pg/mL) Mean (SD) 7.6 (5.71) 6.4 (7.66) 18.4 (6.24) 19.9 (14.30)Median 4.9 5.0 18.4 18.9 Minimum, 1.9, −6.3, 7.3, −11.6, Maximum 22.621.0 29.9 48.3 AUC_(0-6 Weeks) (g · d/mL) Mean (SD) 11.5 (112.97) 100.6(185.38) 249.9 (198.83) 371.1 (290.81) Median 16.2 23.0 298.7 352.2Minimum, −267.1, −145.4, −191.7, −5.8, Maximum 219.8 452.3 563.6 1235.8t_(max) (d) Mean (SD) 24.4 (15.55) 16.8 (16.09) 26.4 (11.52) 25.5(13.88) Median 23.0 12.0 23.0 23.00 Minimum, 2.0, 1.0, 8.0, 1.0, Maximum45.0 44.0 44.0 44.0

The stabilized controlled release formulations of 25-hydroxyvitamin D₃increased serum total 25-hydroxyvitamin D levels to ≥30 ng/mL insignificantly greater number of subjects in all active groups comparedto placebo. Similarly, the stabilized formulations significantlydecreased mean plasma iPTH from baseline in all dose groups compared toplacebo.

Daily administration of 25-hydroxyvitamin D₃ in a stabilized controlledrelease formulation increased mean serum total 25-hydroxyvitamin D inproportion to the dose administered. The lowest administered dose (30μg) increased serum total 25-hydroxyvitamin D at end of treatment by15.6±1.7 (SE) ng/mL from pre-treatment baseline (21.7±1.8 ng/mL) and thehighest dose (90 μg) increased serum total 25-hydroxyvitamin D by61.1±6.1 ng/mL from 21.8±1.2 ng/mL. In contrast, a decrease at end oftreatment of 1.2±0.7 ng/mL was observed in the combined placebo groups.Differences between the treatment and placebo groups were significantfor all three dose levels studied (p<0.0001). The mean serum25-hydroxyvitamin D level in the 30 μg dose group at the end oftreatment was 37.3±1.8 ng/mL, (slightly higher than K/DOQI-specifiedminimum adequate level of 30 ng/mL), indicating that 30 μg was theminimum effective dose.

The percentage of treated subjects achieving serum total25-hydroxyvitamin D levels of ≥30 ng/mL at end of treatment was 92.3%,100.0% and 100.0% in the 30 μg, 60 μg and 90 μg dose groups comparedwith 0% in the placebo group. These differences in response ratesbetween active and placebo treatment were all significant (p<0.001).

Mean plasma iPTH decreased at end of treatment in proportion to theadministered dose of 25-hydroxyvitamin D₃. The lowest administered dose(30 μg) decreased iPTH by 20.2±5.8 (SE) % from pre-treatment baseline,and the highest dose (90 μg) decreased iPTH by 35.9±4.2%. An increase of17.2±7.8% was observed at end of treatment in the combined placebogroups. Differences between groups receiving 25-hydroxyvitamin D₃ andplacebo were significant for all three dose levels studied (p<0.005) andthey compared favorably with differences observed with longer treatmentin placebo-controlled studies with the more potent and calcemic oralvitamin D hormone replacement therapies (e.g., doxercalciferol,paricalcitol and calcitriol).

Percentages of subjects receiving 25-hydroxyvitamin D₃ who achievedconfirmed reductions (i.e., two consecutive measurements) in iPTH of atleast 20% or 30% from pre-treatment baseline at EOT increased with dosethrough 60 μg. Similar response rates were observed in the 60 and 90 μgtreatment groups, indicating that no further benefit in iPTH loweringwas observed in this study for the 90 μg dose. Response rates for aconfirmed 20% reduction in iPTH were 38.5%, 70.6% and 76.5% for the 30μg, 60 μg and 90 μg dose groups respectively compared with 9.7% in thecombined placebo group. Differences in the observed response rates for a20% reduction were significant only for the 60 μg and 90 μg dose groups(p<0.005) and for a 30% reduction were significant in all three dosegroups (p<0.05). The data supported the conclusion that 30 μg per day of25-hydroxyvitamin D₃ in a stabilized controlled release formulation isthe minimum effective dose.

The stabilized formulations of 25-hydroxyvitamin D₃ had no clinicallysignificant effect on corrected albumin-corrected serum calcium, serumphosphorus and urinary calcium excretion. There were no adverse effectson serum calcium or serum phosphorus or urine calcium during the 6-weektreatment period.

Pharmacokinetic analyses revealed that the stabilized formulations of25-hydroxyvitamin D₃ increased 25-hydroxyvitamin D₃ exposure over 6weeks (AUC and C_(max)) in a dose proportional manner across the threedose groups with no difference in t_(1/2). Following 6 weeks ofadministration, the three treatment groups had not quite reached steadystate. However, steady state modeling demonstrated that steady statewould have been achieved by 7-9 weeks in all dose groups.

Data from this study clearly demonstrated that stabilized controlledrelease formulations of 25-hydroxyvitamin D₃ were effective in elevatingserum total 25-hydroxyvitamin D to the minimum adequate level of 30ng/mL and lowering plasma iPTH. The study also showed that stabilizedformulations of 25-hydroxyvitamin D₃ had no clinically meaningful impacton serum calcium or phosphorous at the doses investigated.

Example 4: Pharmacokinetic and Pharmacodynamic Profile ofModified-Release Calcifediol in CKD Subjects with SecondaryHyperparathyroidism and Vitamin D Insufficiency

A multi-center, randomized, double blind, placebo-controlled, repeatdose, safety, efficacy and PK/PD study of stabilized, sustained release25-hydroxyvitamin D₃ (calcifediol, 25D₃) capsules was conducted in 2cohorts of subjects. Male and female subjects aged 18 to 85 years withstage 3 CKD (eGFR of 25-70 mL/min/1.73 m²), vitamin D insufficiency(serum 25-hydroxyvitamin D ≥10 and ≤29 ng/mL), SHPT (plasma iPTH >70μg/mL) and not requiring regular hemodialysis, were recruited for thisstudy. Eligible subjects in the first cohort were randomized into 3treatment groups in a 1:1:1 ratio: 2 groups received the capsules atdaily oral doses of 60 or 90 μg, respectively, and 1 group received amatching placebo capsule. Subjects in cohort 2 were randomized into 2treatment groups in a 1:1 ratio: 1 group received 30 μg capsules dailyand the other received placebo. Subjects in each cohort completed 6weeks of treatment and entered a 6 week follow-up period, during whichPK and PD samples were collected weekly. Serum calcium (Ca), phosphorus(P), 25D₃, total 1,25-dihydroxyvitamin D (1,25D) and plasma iPTH weremonitored weekly during 6 weeks of treatment and 6 weeks of follow-up.ANCOVA models examined the association of 25D₃ exposure with change frombaseline for 1,25D and iPTH. Covariates included were baseline eGFR,body weight and height, gender, age, race, diabetic status, and baselineconcentration of 1,25D or iPTH.

FIG. 4 shows the resulting mean baseline-adjusted calcifediolconcentrations by treatment group (PK population). Mean levels of serumcalcifediol increased gradually and in a dose-proportional manner andbegan to approach steady-state by 6 weeks. After 6 weeks of follow-up,levels decreased but remained above baseline in all active-treatedgroups.

FIG. 5 shows the resulting summary baseline-adjusted PK parameters forcalcifediol concentrations by treatment group (PK population).

FIG. 6 shows the resulting mean baseline-adjusted serum1,25-dihydroxyvitamin D levels during the 6-week treatment (PKpopulation). Mean baseline-adjusted serum total 1,25 dihydroxyvitamin Dlevels increased over time in those subjects administered the activecapsules, compared to those subjects administered placebo.

FIG. 7 shows a summary of the resulting baseline-adjusted repeat-dose PKparameters for serum 1,25-dihydroxyvitamin D by treatment group (PKpopulation).

FIG. 8 shows the resulting mean percent of baseline in plasma iPTHlevels during the 6-week treatment (PK population). The active capsulessignificantly decreased mean plasma iPTH from baseline by 21%, 33% and39% in all dose groups (30, 60 and 90 μg, respectively) compared to a17% increase in the combined placebo group.

FIG. 9 shows a summary of the resulting baseline-adjusted repeat-dose PKparameters for plasma iPTH by treatment group (PK population).

FIGS. 10 and 11 show the percent change from baseline at EOT for plasmaiPTH relative to baseline-adjusted calcifediol and 1,25-dihydroxyvitaminD exposure (AUC_(0-6wk)) in the PK Population. Percent reductions inplasma iPTH from baseline to EOT increased as serum calcifediol andtotal 1,25 dihydroxyvitamin D exposures during treatment (expressed asbaseline-adjusted AUC_(0-6wk)) increased.

The stabilized, sustained release 25-hydroxyvitamin D₃ capsulesnormalized 25D levels in the majority of subjects and significantlyreduced iPTH in all dose groups (30, 60 and 90 μg). The stabilized,sustained release 25-hydroxyvitamin D₃ capsules increased serum 25D₃ andserum 1,25D levels gradually with dose-dependent increases in exposure.Both 25D₃ and total 1,25D exposure were significantly and inverselyassociated with change from baseline for plasma iPTH. Only eGFR was asignificant covariate in both models. These findings demonstrate thatthe stabilized, sustained release 25-hydroxyvitamin D₃ capsules reliablynormalized 25D levels, increased serum 1,25D levels and suppressedelevated plasma iPTH without clinically meaningful effects on serum Caand P at the doses investigated.

The foregoing description is given for clearness of understanding only,and no unnecessary limitations should be understood therefrom, asmodifications within the scope of the invention may be apparent to thosehaving ordinary skill in the art.

Throughout this specification and the claims which follow, unless thecontext requires otherwise, the word “comprise” and variations such as“comprises” and “comprising” will be understood to imply the inclusionof a stated integer or step or group of integers or steps but not theexclusion of any other integer or step or group of integers or steps.

Throughout the specification, where compositions are described asincluding components or materials, it is contemplated that thecompositions can also consist essentially of, or consist of, anycombination of the recited components or materials, unless describedotherwise. Likewise, where methods are described as including particularsteps, it is contemplated that the methods can also consist essentiallyof, or consist of, any combination of the recited steps, unlessdescribed otherwise. The invention illustratively disclosed hereinsuitably may be practiced in the absence of any element or step which isnot specifically disclosed herein.

The practice of a method disclosed herein, and individual steps thereof,can be performed manually and/or with the aid of electronic equipment.Although processes have been described with reference to particularembodiments, a person of ordinary skill in the art will readilyappreciate that other ways of performing the acts associated with themethods may be used. For example, the order of various of the steps maybe changed without departing from the scope or spirit of the method,unless described otherwise. In addition, some of the individual stepscan be combined, omitted, or further subdivided into additional steps.

All patents, publications and references cited herein are hereby fullyincorporated by reference. In case of conflict between the presentdisclosure and incorporated patents, publications and references, thepresent disclosure should control.

Embodiments contemplated in view of the foregoing description includethose described in the following numbered paragraphs.

1. A controlled release formulation of a vitamin D compound comprisingone or both of 25-hydroxyvitamin D₂ and 25-hydroxyvitamin D₃, theformulation comprising a matrix that releasably binds and controllablyreleases the vitamin D compound, the matrix comprising a cellulosederivative.

2. A stabilized formulation for controlled release of a vitamin Dcompound in the gastrointestinal tract of a subject which ingests theformulation, the formulation comprising a mixture of:

one or both of 25-hydroxyvitamin D₂ and 25-hydroxyvitamin D₃;

and an effective amount of a stabilizing agent, which is optionally acellulosic compound, to maintain a difference of less than 30% betweenthe amount of vitamin D compound released at any given time point afterfour hours during in vitro dissolution after two months exposure tostorage conditions of 25° C. and 60% relative humidity and the amountreleased at the same dissolution time point during in vitro dissolutionconducted prior to exposing the formulation to the storage conditions.

3. A stabilized formulation for controlled release of a vitamin Dcompound, said formulation comprising a mixture of:

one or both of 25-hydroxyvitamin D₂ and 25-hydroxyvitamin D₃;

a wax matrix; and

a stabilizing agent, which is optionally a cellulosic compound.

4. A stabilized formulation for controlled release of a vitamin Dcompound in the gastrointestinal tract of a subject which ingests theformulation, said formulation comprising a mixture of:

one or both of 25-hydroxyvitamin D₂ and 25-hydroxyvitamin D₃;

a wax matrix; and

a stabilizing agent, which is optionally a cellulosic stabilizing agent.

5. A stabilized formulation for controlled release of a vitamin D, saidformulation comprising a mixture of:

one or both of 25-hydroxyvitamin D₂ and 25-hydroxyvitamin D₃;

a wax matrix; and

an effective amount of a stabilizing agent, optionally a cellulosiccompound, to maintain a difference of less than 30% between the amountof vitamin D compound released at any given time point after four hoursduring in vitro dissolution testing after exposure for two months tostorage conditions of 25° C. and 60% relative humidity and the amountreleased at the same dissolution time point during in vitro dissolutionconducted prior to exposing the formulation to the storage conditions.

6. A stabilized formulation for controlled release of a vitamin Dcompound in the gastrointestinal tract of a subject which ingests theformulation, said formulation comprising a mixture of:

an active-loaded wax matrix comprising one or both of 25-hydroxyvitaminD₂ and 25-hydroxyvitamin D₃; and

a cellulosic stabilizing agent;

wherein the formulation releases an amount of 25-hydroxyvitamin D duringin vitro dissolution after two months exposure to storage of 25° C. and60% relative humidity that varies at any given dissolution time pointcompared to the amount released at the same dissolution time pointduring in vitro dissolution conducted prior to exposing the formulationto the storage conditions by 30% or less in the absence of thecellulosic stabilizing agent.

7. In a formulation for controlled release of a vitamin D compound inthe gastrointestinal tract of a subject which ingests the formulation,the improvement comprising admixing a cellulosic stabilizing agent intothe formulation.

8. In a formulation for controlled release of a vitamin D compound inthe gastrointestinal tract of a subject which ingests the formulation,the improvement comprising an effective amount of a cellulosic compoundadmixed in the formulation to maintain a change of less than 30% in theamount of 25-hydroxyvitamin D released during in vitro dissolution afterexposure to storage conditions of at least one month at 25° C. and 60%relative humidity at any given dissolution time point after four hourscompared to the amount released at the same dissolution time pointduring in vitro dissolution conducted prior to exposing the formulationto the storage conditions.

9. The formulation according to any of the preceding paragraphs, whereinthe cellulosic compound or cellulosic stabilizing agent comprises acellulose ether.

10. The formulation according to paragraph 9, wherein the celluloseether is selected from the group consisting of methylcellulose, hydroxylpropyl methylcellulose, hydroxyl ethyl methylcellulose, hydroxyl ethylcellulose, and hydroxyl propyl cellulose.

11. The formulation according to paragraph 9, wherein the cellulosiccompound or cellulosic stabilizing agent is hydroxyl propylmethylcellulose.

12. The formulation according to any of the preceding paragraphs,wherein the formulation releases an amount of 25-hydroxyvitamin D duringin vitro dissolution after exposure to storage conditions for 2 monthsat 25° C. and 60% relative humidity that varies at any given dissolutiontime point after four hours compared to the amount released at the samedissolution time point during in vitro dissolution conducted prior toexposing the formulation to the storage conditions, by 30% or less.

13. The formulation according to any of the preceding paragraphs,wherein the formulation releases an amount of 25-hydroxyvitamin D duringin vitro dissolution after exposure to storage conditions for one monthat 40° C. and 75% relative humidity that varies at any given dissolutiontime point after four hours compared to the amount released at the samedissolution time point during in vitro dissolution conducted prior toexposing the formulation to the storage conditions, by 30% or less.

14. The formulation according to any one of the preceding paragraphs,wherein the matrix comprises a wax matrix comprising a controlledrelease agent, an emulsifier, and an absorption enhancer.

15. The formulation according to paragraph 14, wherein the controlledrelease agent comprises paraffin.

16. The formulation according to paragraph 14 or 15, wherein theemulsifier has a HLB value less than 7.

17. The formulation according to paragraph 16, wherein the emulsifiercomprises glycerol monostearate.

18. The formulation according to any one of paragraphs 14 to 17, whereinthe absorption enhancer has a HLB value in a range of about 13 to about18.

19. The formulation according to paragraph 18, wherein the absorptionenhancer is a mixture of lauroyl macrogolglycerides and lauroylpolyoxylglycerides.

20. The formulation according to any of the preceding paragraphs,wherein the vitamin D compound comprises 25-hydroxyvitamin D₃.

21. The formulation according to any of the preceding paragraphs,further comprising an oily vehicle.

22. The formulation according to paragraph 21, wherein the oily vehiclecomprises mineral oil.

23. The formulation according to paragraph 22, wherein the formulationcomprises about 20 wt % paraffin, about 20 wt % to about 25 wt %glycerol monostearate, about 10 wt % a mixture of lauroylmacrogolglycerides and lauroyl polyoxylglycerides, about 30 wt % toabout 35 wt % mineral oil, and about 10 wt % to about 15 wt % hydroxylpropyl methylcellulose.

24. A formulation according to any one of the preceding paragraphs,wherein the formulation comprises glycerol monostearate.

25. A formulation according to any one of the preceding paragraphs,wherein the formulation comprises one or more polyglycolized glycerides.

26. A sustained release dosage form in the form of a capsule, tablet,sachet, dragee, or suppository comprising a formulation according to anyone of the preceding paragraphs.

27. The dosage form according to paragraph 26, comprising a capsule ortablet.

28. The dosage form according to paragraph 27, comprising a capsule.

29. The dosage form according to paragraph 26, comprising an oralcapsule, tablet, sachet, dragee.

30. A stabilized dosage form according to any one of the precedingparagraphs characterized by a dissolution profile providing a release ofvitamin D compound of

less than 30% at 2 hours;

greater than 45% at 6 hours; and

greater than 80% at 12 hours.

31. The stabilized dosage form according to paragraph 26, wherein therelease of vitamin D compound at 6 hours is less than 60%.

32. A stabilized sustained release oral dosage form comprising a vitaminD compound characterized by an in vitro dissolution profile providingrelease of vitamin D compound of

less than 30% at 100 to 140 minutes;

greater than 45% at 5 to 7 hours; and

greater than 80% at 11 to 13 hours.

33. The dosage form of paragraph 32, wherein the release of vitamin Dcompound is

less than 30% at 2 hours;

greater than 45% at 6 hours; and

greater than 80% at 12 hours.

34. The dosage form according to paragraph 32 or 33, wherein the releaseof vitamin D compound at 5 to 7 hours is less than 60%.

35. The dosage form according to paragraph 34, wherein the release ofvitamin D compound at 6 hours is less than 60%.

36. A stabilized sustained release oral dosage form comprising a vitaminD compound characterized by an in vitro dissolution profile providingrelease of vitamin D compound of

about 20% to about 40% at 2 hours;

at least 35% at 6 hours; and

at least 70% at 12 hours.

37. The dosage form of paragraph 36, wherein the release of vitamin Dcompound is about 25% to about 35% at 2 hours;

at least 40% at 6 hours; and

at least 75% at 12 hours.

38. The dosage form of paragraph 36 or 37, wherein the release ofvitamin D compound is 75% or less at 6 hours.

39. The dosage form of paragraph 38, wherein the release of vitamin Dcompound is 65% or less at 6 hours.

40. The dosage form of paragraph 39, wherein the release of vitamin Dcompound is 60% or less at 6 hours.

41. A stabilized sustained release dosage form comprising a vitamin Dcompound characterized by a t_(max) following administration of thedosage form to a human patient, of at least 4 hours.

42. The dosage form of paragraph 41, wherein the t_(max) is at least 8hours.

43. The dosage form of paragraph 42, wherein the t_(max) is at least 12hours.

44. The dosage form of paragraph 43, wherein the t_(max) is at least 18hours.

45. The dosage form of paragraph 44, wherein the t_(max) is at least 20hours.

46. The dosage form of paragraph 45, wherein the t_(max) is at least 24hours.

47. The dosage form of paragraph 46, wherein the t_(max) is at least 28hours.

48. The dosage form of paragraph 41, wherein the t_(max) is in a rangeof 4 to 96 hours.

49. The dosage form of paragraph 48, wherein the t_(max) is in a rangeof 18 to 30 hours.

50. The dosage form of paragraph 49, wherein the t_(max) is in a rangeof 13 to 28 hours.

51. The dosage form of paragraph 50, wherein the t_(max) is about 28hours.

52. A stabilized sustained release dosage form comprising a25-hydroxyvitamin D compound, the dosage form characterized by providinga baseline-adjusted C_(max) per microgram of 25-hydroxyvitamin D in arange of about 0.0133 ng/mL to about 0.04 ng/mL when administered to anadult human.

53. A method of administering a stabilized sustained release dosage formcomprising a 25-hydroxyvitamin D compound to a human patient, comprisingadministering an effective amount of the dosage form to the patient toprovide a baseline-adjusted Cmax of at least about 0.2 ng/mL and lessthan 110 ng/mL.

54. The method of paragraph 53, comprising administering an effectiveamount of the dosage form to provide a baseline-adjusted Cmax in a rangeof about 0.2 to about 24 ng/mL.

55. A method of administering a stabilized sustained release dosage formcomprising a 25-hydroxyvitamin D compound to a human patient, comprisingadministering an effective amount of the dosage form to the patient toprovide a baseline-adjusted AUC_(0-inf) of at least 52 ng*h/mL and lessthan 34500 ng*h/mL.

56. The method of paragraph 55, comprising administering an effectiveamount of the dosage form to the patient to provide a baseline-adjustedAUC_(0-inf) in a range of about 52 ng*h/mL to about 12,000 ng*h/mL.

57. A method of vitamin D supplementation comprising administering to asubject in need thereof a formulation or dosage form according to anyone of the preceding paragraphs.

58. A method of treatment or prophylaxis of a vitamin-D responsivedisease in a subject comprising administering to the subject aformulation or dosage form according to any one of the precedingparagraphs.

59. The method of paragraph 58, wherein the disease is selected fromcancer (e.g., breast, lung, skin, melanoma, colon, colorectal, rectal,prostate and bone cancer), autoimmune diseases, for example, type Idiabetes, multiple sclerosis, rheumatoid arthritis, polymyositis,dermatomyositis, scleroderma, fibrosis, Grave's disease, Hashimoto'sdisease, acute or chronic transplant rejection, acute or chronic graftversus host disease, inflammatory bowel disease, Crohn's disease,systemic lupus erythematosis, Sjogren's Syndrome, eczema and psoriasis,dermatitis, including atopic dermatitis, contact dermatitis, allergicdermatitis and/or chronic dermatitis, inflammatory diseases, forexample, asthma, chronic obstructive pulmonary disease, polycystickidney disease, polycystic ovary syndrome, pancreatitis, nephritis,hepatitis, and/or infection, hypertension, cardiovascular diseases, forexample, subjects with atherosclerosis, arteriosclerosis, coronaryartery disease, cerebrovascular disease, peripheral vascular disease,myocardial infarction, myocardial ischemia, cerebral ischemia, stroke,congestive heart failure, cardiomyopathy, obesity or other weightdisorders, lipid disorders (e.g. hyperlipidemia, dyslipidemia includingassociated diabetic dyslipidemia and mixed dyslipidemiahypoalphalipoproteinemia, hypertriglyceridemia, hypercholesterolemia,and low HDL (high density lipoprotein)), metabolic disorders (e.g.Metabolic Syndrome, Type II diabetes mellitus, Type I diabetes mellitus,hyperinsulinemia, impaired glucose tolerance, insulin resistance,diabetic complication including neuropathy, nephropathy, osteoporosis,retinopathy, diabetic foot ulcer and cataracts), and/or thrombosis.

60. The method of paragraph 59, wherein the disease is selected from (i)in the parathyroid—hypoparathyroidism, Pseudohypo-parathyroidism,secondary hyperparathyroidism; (ii) in the pancreas—diabetes; (iii) inthe thyroid—medullary carcinoma; (iv) in the skin—psoriasis; woundhealing; (v) in the lung—sarcoidosis and tuberculosis; (vi) in thekidney—chronic kidney disease, hypophosphatemic VDRR, vitamin Ddependent rickets; (vii) in the bone—anticonvulsant treatment,fibrogenisis imperfecta ossium, osteitis fibrosa cystica, osteomalacia,osteoporosis, osteopenia, osteosclerosis, renal osteodytrophy, rickets;(viii) in the intestine—glucocorticoid antagonism, idopathichypercalcemia, malabsorption syndrome, steatorrhea, tropical sprue; and(ix) autoimmune disorders.

61. The method of paragraph 60, wherein the disease is selected fromcancer, dermatological disorders (for example psoriasis), parathyroiddisorders (for example hyperparathyroidism and secondaryhyperparathyroidism), bone disorders (for example osteoporosis) andautoimmune disorders.

62. The method of paragraph 61, wherein the disease is secondaryhyperparathyroidism.

63. The method of paragraph 62, wherein the subject has chronic kidneydisease (CKD).

64. The method of paragraph 63, wherein the CKD is Stage 3 or 4.

65. The method of paragraph 64, wherein the patient is vitamin-Ddeficient.

66. The method of any one of the preceding paragraphs, wherein thepatient is human.

67. The method of paragraph 66, wherein the human is an adult human.

68. A composition as substantially herein described.

What is claimed:
 1. A stabilized formulation for extended release of avitamin D compound in the gastrointestinal tract of a subject whichingests the formulation, the formulation comprising a mixture of: one orboth of 25-hydroxyvitamin D₂ and 25-hydroxyvitamin D₃ in an extendedrelease lipophilic matrix, the lipophilic matrix further comprising acellulosic stabilizing agent to stabilize the matrix against changes indissolution release characteristics over time; wherein the formulationis characterized by providing a mean t_(max) of the vitamin D compoundin adult humans following an initial, single dose, in a range of 4 to 96hours.
 2. The stabilized formulation of claim 1, wherein the meant_(max) is at least 18 hours.
 3. The stabilized formulation of claim 2,wherein the mean t_(max) is in a range of 18 to 30 hours.
 4. Thestabilized formulation of claim 2, wherein the mean t_(max) is at least20 hours.
 5. The stabilized formulation of claim 2, wherein the meant_(max) is at least 24 hours.
 6. The stabilized formulation of claim 2,wherein the mean t_(max) is at least 28 hours.
 7. The stabilizedformulation of claim 1, wherein the lipophilic matrix comprises a wax.8. The stabilized formulation of claim 7, wherein the lipophilic matrixcomprises paraffin wax.
 9. The stabilized formulation according to claim1, further comprising an emulsifier.
 10. The stabilized formulationaccording to claim 9, wherein the emulsifier has a HLB value less than7.
 11. The stabilized formulation according to claim 10, wherein theemulsifier comprises glycerol monostearate.
 12. The stabilizedformulation according to claim 1, further comprising an absorptionenhancer.
 13. The stabilized formulation according to claim 12, whereinthe absorption enhancer has a HLB value in a range of about 13 to about18.
 14. The stabilized formulation according to claim 12, wherein theabsorption enhancer is a mixture of lauroyl macrogolglycerides andlauroyl polyoxylglycerides.
 15. The stabilized formulation according toclaim 1, further comprising an oily vehicle.
 16. The stabilizedformulation according to claim 15, wherein the oily vehicle comprisesmineral oil.
 17. The stabilized formulation according to claim 1,wherein the lipophilic matrix comprises a wax, an oil, an emulsifier,and an absorption enhancer.
 18. The stabilized formulation according toclaim 1, wherein the formulation comprises about 20 wt % paraffin, about20 wt % to about 25 wt % glycerol monostearate, about 10 wt % a mixtureof lauroyl macrogolglycerides and lauroyl polyoxylglycerides, about 30wt % to about 35 wt % mineral oil, and about 10 wt % to about 15 wt %hydroxyl propyl methylcellulose.
 19. The stabilized formulationaccording to claim 18, wherein the vitamin D compound comprises25-hydroxyvitamin D₃.
 20. The stabilized formulation of claim 1, whereinthe cellulosic compound comprises a cellulose ether.
 21. The stabilizedformulation according to claim 20, wherein the cellulose ether comprisesone or more of methylcellulose, hydroxyl propyl methylcellulose,hydroxyl ethyl methylcellulose, hydroxyl ethyl cellulose, and hydroxylpropyl cellulose.
 22. The stabilized formulation according to claim 21,wherein the cellulosic compound comprises hydroxyl propylmethylcellulose.
 23. An extended release dosage form in the form of acapsule, tablet, sachet, or dragee comprising a stabilized formulationaccording to claim
 1. 24. An extended release capsule for extendedrelease of 25-hydroxyvitamin D₃ in the gastrointestinal tract of asubject which ingests the capsule, the capsule containing25-hydroxyvitamin D₃ in an extended release, wax-based matrix, thematrix comprising paraffin, mineral oil, an emulsifier, and a celluloseether, wherein the formulation is characterized by providing a meant_(max) of the vitamin D compound in adult humans following an initial,single dose, in a range of 18 to 30 hours.
 25. The stabilizedformulation of claim 1, wherein the formulation is further characterizedby providing a mean baseline-adjusted in vivo C_(max) per microgram of25-hydroxyvitamin D administered to humans in a range of about 0.0133ng/mL to about 0.04 ng/mL.
 26. The stabilized formulation of claim 25,wherein the formulation, following storage for 3 months at 25° C. and60% RH, provides a C_(max) within 80% to 125% of the mean of the freshproduct.
 27. The stabilized formulation of claim 1, wherein theformulation is further characterized by a baseline-adjusted AUC_(0-6wk)of the administered vitamin D compound having a mean of about 700ng·d/mL, when administered to adult humans having CKD Stage 3 withsecondary hyperparathyroidism and vitamin D insufficiency, and at dailyoral doses of 30 mcg.
 28. The stabilized formulation of claim 27,wherein the formulation comprises 30 mcg 25-hydroxyvitamin D₃.
 29. Amethod of administering a stabilized formulation for extended release ofa vitamin D compound in the gastrointestinal tract of a subject whichingests the formulation, the formulation comprising a mixture of one orboth of 25-hydroxyvitamin D₂ and 25-hydroxyvitamin D₃ in an extendedrelease lipophilic matrix, the lipophilic matrix further comprising acellulosic stabilizing agent to stabilize the matrix against changes indissolution release characteristics over time; comprising administeringan effective amount of the formulation to a human patient to provide abaseline-adjusted AUC_(0-inf) following a dose in a range of 52 ng·h/mLto less than 34,500 ng·h/mL.
 30. The method of claim 29, comprisingadministering an effective amount of the formulation to a human patientto provide a baseline-adjusted AUC_(0-inf) following a dose in a rangeof 52 ng·h/mL to about 12,000 ng·h/mL.